bt474  (ATCC)


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    Structured Review

    ATCC bt474
    Characterization of HER2-positive BC cell lines. ( a ) Lapatinib dose–response curves for HER2-positive BC cell lines. Results are normalized to those of untreated cells and are representative of two independent experiments. ( b ) Western blot analysis of HER2-positive BC cell extracts after immunoprecipitation with anti-HER2 antibodies. Images are representative of two independent experiments. Ratios were calculated for band quantification by Quantity One. ER-positive and HER2-addicted BC cell lines are shown in red and underlined, respectively. ( c ) HER2 mRNA levels as evaluated by qRT-PCR. Data are the mean ± SD of three independent experiments and are relative to <t>BT474</t> HER2 expression levels. ( d ) Western blot analysis of HER2-positive BC cell extracts. Vinculin was used as a loading control. Images are representative of two independent experiments. The names of ER-positive and HER2-addicted BC cell lines are shown in red and underlined, respectively. The asterisks indicate molecules analyzed in a separate gel.
    Bt474, supplied by ATCC, used in various techniques. Bioz Stars score: 97/100, based on 40 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    bt474 - by Bioz Stars, 2022-11
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    Images

    1) Product Images from "HER2 mRNA Levels, Estrogen Receptor Activity and Susceptibility to Trastuzumab in Primary Breast Cancer"

    Article Title: HER2 mRNA Levels, Estrogen Receptor Activity and Susceptibility to Trastuzumab in Primary Breast Cancer

    Journal: Cancers

    doi: 10.3390/cancers14225650

    Characterization of HER2-positive BC cell lines. ( a ) Lapatinib dose–response curves for HER2-positive BC cell lines. Results are normalized to those of untreated cells and are representative of two independent experiments. ( b ) Western blot analysis of HER2-positive BC cell extracts after immunoprecipitation with anti-HER2 antibodies. Images are representative of two independent experiments. Ratios were calculated for band quantification by Quantity One. ER-positive and HER2-addicted BC cell lines are shown in red and underlined, respectively. ( c ) HER2 mRNA levels as evaluated by qRT-PCR. Data are the mean ± SD of three independent experiments and are relative to BT474 HER2 expression levels. ( d ) Western blot analysis of HER2-positive BC cell extracts. Vinculin was used as a loading control. Images are representative of two independent experiments. The names of ER-positive and HER2-addicted BC cell lines are shown in red and underlined, respectively. The asterisks indicate molecules analyzed in a separate gel.
    Figure Legend Snippet: Characterization of HER2-positive BC cell lines. ( a ) Lapatinib dose–response curves for HER2-positive BC cell lines. Results are normalized to those of untreated cells and are representative of two independent experiments. ( b ) Western blot analysis of HER2-positive BC cell extracts after immunoprecipitation with anti-HER2 antibodies. Images are representative of two independent experiments. Ratios were calculated for band quantification by Quantity One. ER-positive and HER2-addicted BC cell lines are shown in red and underlined, respectively. ( c ) HER2 mRNA levels as evaluated by qRT-PCR. Data are the mean ± SD of three independent experiments and are relative to BT474 HER2 expression levels. ( d ) Western blot analysis of HER2-positive BC cell extracts. Vinculin was used as a loading control. Images are representative of two independent experiments. The names of ER-positive and HER2-addicted BC cell lines are shown in red and underlined, respectively. The asterisks indicate molecules analyzed in a separate gel.

    Techniques Used: Western Blot, Immunoprecipitation, Quantitative RT-PCR, Expressing

    2) Product Images from "Sabizabulin, a Potent Orally Bioavailable Colchicine Binding Site Agent, Suppresses HER2+ Breast Cancer and Metastasis"

    Article Title: Sabizabulin, a Potent Orally Bioavailable Colchicine Binding Site Agent, Suppresses HER2+ Breast Cancer and Metastasis

    Journal: Cancers

    doi: 10.3390/cancers14215336

    Efficacy of the combination of either paclitaxel or sabizabulin with lapatinib on cell growth inhibition and wound healing. ( A ) Lapatinib sensitivity of BT474, SKBR3, AU565 and JIMT HER2+ cell lines was measured by the MTS assay over a range of doses, data enumerated as a percentage of viable cells, and plotted on log (nM) scale, confirming that JIMT cells are lapatinib-resistant. ( B , C ) The MTS assay was used to determine the effect of pairing lapatinib (Lap) with paclitaxel (left panels) or sabizabulin (right panels) in AU565 ( B ) or JIMT ( C ) cells by the isobole method; the effect of the anti-tubulin targeting agent alone at the IC 30 dose is shown as a dashed line. ( B , C ) Data are plotted as the % maximal proliferation on a log scale (M = molar) after normalization to the untreated control as described in the Materials and Methods. Assays in ( A – C ) are representative of 3 to 4 biological replicates. ( D ) Wound healing was assayed by measuring wound width over time after drug added using the IncuCyte S3 instrument. JIMT cells were treated with either lapatinib (Lap), sabizabulin or paclitaxel alone, or in combination with 100 nM of lapatinib (n = 8 wells/cell line/treatment). Data are representative of two biological replicate experiments. ** p
    Figure Legend Snippet: Efficacy of the combination of either paclitaxel or sabizabulin with lapatinib on cell growth inhibition and wound healing. ( A ) Lapatinib sensitivity of BT474, SKBR3, AU565 and JIMT HER2+ cell lines was measured by the MTS assay over a range of doses, data enumerated as a percentage of viable cells, and plotted on log (nM) scale, confirming that JIMT cells are lapatinib-resistant. ( B , C ) The MTS assay was used to determine the effect of pairing lapatinib (Lap) with paclitaxel (left panels) or sabizabulin (right panels) in AU565 ( B ) or JIMT ( C ) cells by the isobole method; the effect of the anti-tubulin targeting agent alone at the IC 30 dose is shown as a dashed line. ( B , C ) Data are plotted as the % maximal proliferation on a log scale (M = molar) after normalization to the untreated control as described in the Materials and Methods. Assays in ( A – C ) are representative of 3 to 4 biological replicates. ( D ) Wound healing was assayed by measuring wound width over time after drug added using the IncuCyte S3 instrument. JIMT cells were treated with either lapatinib (Lap), sabizabulin or paclitaxel alone, or in combination with 100 nM of lapatinib (n = 8 wells/cell line/treatment). Data are representative of two biological replicate experiments. ** p

    Techniques Used: Inhibition, MTS Assay

    Effect of sabizabulin, colchicine, or paclitaxel in BT474 and SKBR3 HER2+ cell lines. ( A ) Chemical structure of sabizabulin. ( B ) Levels of HER2 in breast cancer cell lines were determined by western blotting; β-actin serves as a loading control. Signal intensity was evaluated by ImageJ densitometry with the AU565 sample set to 1.0. ( C ) Proliferation assay over time (120 h) using BT474 and SKBR3 cells to determine doubling times by confluence (%) using the IncuCyte S3 instrument (5000 cells/96-well; n = 4 wells/cell line). ( D ) The growth inhibition effect of sabizabulin relative to colchicine or paclitaxel was determined by MTS assay after 72 h or 96 h (SKBR3 or BT474, respectively) over a range of concentrations (0.3 nM to 3.0 µM, on a log (nM) scale) and expressed as cell viability (%)representative response curves of three independent biological replicates are shown. ( E , F ) Growth inhibition assays for BT474 cells ( E ) or SKBR3 cells ( F ) treated with paclitaxel or sabizabulin at increasing concentrations as measured by confluence (%) over time (hrs). ( G , H ) Evaluation of time- and concentration-dependent cytotoxicity for BT474 ( G ) and SKBR3 ( H ) cells as enumerated by Cytotox Green reagent counts (Green Object per image) over time (hrs).All p -values are shown as pairwise comparisons to the vehicle (control) group; * p
    Figure Legend Snippet: Effect of sabizabulin, colchicine, or paclitaxel in BT474 and SKBR3 HER2+ cell lines. ( A ) Chemical structure of sabizabulin. ( B ) Levels of HER2 in breast cancer cell lines were determined by western blotting; β-actin serves as a loading control. Signal intensity was evaluated by ImageJ densitometry with the AU565 sample set to 1.0. ( C ) Proliferation assay over time (120 h) using BT474 and SKBR3 cells to determine doubling times by confluence (%) using the IncuCyte S3 instrument (5000 cells/96-well; n = 4 wells/cell line). ( D ) The growth inhibition effect of sabizabulin relative to colchicine or paclitaxel was determined by MTS assay after 72 h or 96 h (SKBR3 or BT474, respectively) over a range of concentrations (0.3 nM to 3.0 µM, on a log (nM) scale) and expressed as cell viability (%)representative response curves of three independent biological replicates are shown. ( E , F ) Growth inhibition assays for BT474 cells ( E ) or SKBR3 cells ( F ) treated with paclitaxel or sabizabulin at increasing concentrations as measured by confluence (%) over time (hrs). ( G , H ) Evaluation of time- and concentration-dependent cytotoxicity for BT474 ( G ) and SKBR3 ( H ) cells as enumerated by Cytotox Green reagent counts (Green Object per image) over time (hrs).All p -values are shown as pairwise comparisons to the vehicle (control) group; * p

    Techniques Used: Western Blot, Proliferation Assay, Inhibition, MTS Assay, Concentration Assay

    Sabizabulin inhibits tumor growth and induces apoptosis in orthotopic BT474 xenografts. ( A ) Mean tumor volume (mm 3 ) was measured over the course of treatments (days on therapy): vehicle (IP, 3×/week, n = 19 total tumors), paclitaxel (10 mg/kg, IP, 3×/week, n = 17 total tumors), or sabizabulin (17 mg/kg, PO, 3×/week, n = 21 total tumors). Mean tumor wet weight ( B ) and mean ex vivo measured tumor volume ( C ) are shown at experiment endpoint (33 days). ( A – C ) The mean values are also indicated above each treatment group. All p -values were calculated as pairwise comparisons relative to the vehicle control group at 33 days; ** p
    Figure Legend Snippet: Sabizabulin inhibits tumor growth and induces apoptosis in orthotopic BT474 xenografts. ( A ) Mean tumor volume (mm 3 ) was measured over the course of treatments (days on therapy): vehicle (IP, 3×/week, n = 19 total tumors), paclitaxel (10 mg/kg, IP, 3×/week, n = 17 total tumors), or sabizabulin (17 mg/kg, PO, 3×/week, n = 21 total tumors). Mean tumor wet weight ( B ) and mean ex vivo measured tumor volume ( C ) are shown at experiment endpoint (33 days). ( A – C ) The mean values are also indicated above each treatment group. All p -values were calculated as pairwise comparisons relative to the vehicle control group at 33 days; ** p

    Techniques Used: Ex Vivo

    Sabizabulin induces apoptosis in BT474 and SKBR3 cells in a concentration-dependent manner. ( A ) A panel of apoptotic markers was profiled by western blotting after 24 h of treatment with colchicine or paclitaxel (100 nM) or sabizabulin (10, 20, 50, or 100 nM) in BT474 and SKBR3 cells. Signal from cleaved-PARP (anti-cleaved-PARP, CST 5625, 1:1000), phosphorylated BCL2 (pBCL2; anti-pBCL2, 2827 CST 1:1000) and cleaved caspase-3 (CST 9661, 1:500) was first normalized to β-actin (CST 3700, 1:20,000) by ImageJ densitometry, with the 100 nM sabizabulin-treated lane set to 1.0. ( B ) Cleaved-PARP and cleaved-caspase-3 levels in both HER2+ cell lines were determined by western blotting after treatment with 100 nM of sabizabulin for 12, 24, 48, and 72 h after normalization to β-actin. Data are representative of three independent biological replicate assays.
    Figure Legend Snippet: Sabizabulin induces apoptosis in BT474 and SKBR3 cells in a concentration-dependent manner. ( A ) A panel of apoptotic markers was profiled by western blotting after 24 h of treatment with colchicine or paclitaxel (100 nM) or sabizabulin (10, 20, 50, or 100 nM) in BT474 and SKBR3 cells. Signal from cleaved-PARP (anti-cleaved-PARP, CST 5625, 1:1000), phosphorylated BCL2 (pBCL2; anti-pBCL2, 2827 CST 1:1000) and cleaved caspase-3 (CST 9661, 1:500) was first normalized to β-actin (CST 3700, 1:20,000) by ImageJ densitometry, with the 100 nM sabizabulin-treated lane set to 1.0. ( B ) Cleaved-PARP and cleaved-caspase-3 levels in both HER2+ cell lines were determined by western blotting after treatment with 100 nM of sabizabulin for 12, 24, 48, and 72 h after normalization to β-actin. Data are representative of three independent biological replicate assays.

    Techniques Used: Concentration Assay, Western Blot

    3) Product Images from "Upregulation of Receptor Tyrosine Kinase Activity and Stemness as Resistance Mechanisms to Akt Inhibitors in Breast Cancer"

    Article Title: Upregulation of Receptor Tyrosine Kinase Activity and Stemness as Resistance Mechanisms to Akt Inhibitors in Breast Cancer

    Journal: Cancers

    doi: 10.3390/cancers14205006

    RNA sequencing reveals upregulation of ID4 in Akt-inhibitor-resistant cells. ( A ) Left: Venn diagram of distribution of differentially expressed genes in resistant lines. S1, T47D parental. S2, T47D MK0.2-5. S3, T47D MK5. S4, BT474 parental. S5, BT474 MK0.2-2. S6, BT474 GDC1-2. Right: Heatmap displaying genes (142) in which expression is changed (logFC > 1 or logFC
    Figure Legend Snippet: RNA sequencing reveals upregulation of ID4 in Akt-inhibitor-resistant cells. ( A ) Left: Venn diagram of distribution of differentially expressed genes in resistant lines. S1, T47D parental. S2, T47D MK0.2-5. S3, T47D MK5. S4, BT474 parental. S5, BT474 MK0.2-2. S6, BT474 GDC1-2. Right: Heatmap displaying genes (142) in which expression is changed (logFC > 1 or logFC

    Techniques Used: RNA Sequencing Assay, Expressing

    Upregulation of CSC phenotypes in Akt-inhibitor-resistant tumor cells. ( A ) BT474 parental and Akt-inhibitor-resistant cells were seeded to plates in the absence of Akt inhibitor for 48 h. Cells were then treated with MK2206 or GDC0068 for 1 h. Whole-cell lysates were subjected to immunoblotting. * Resistant cells were cultured in the absence of Akt inhibitor for 3.5 weeks, before Western blot analysis. ( B ) Mammosphere forming assays of parental and resistant cells. BT474 cells were seeded for first-generation (1°) mammosphere formation assay. T47D cells were seeded for first generation of mammosphere formation assay, followed by second and then third generation (3°). Mammosphere forming efficiency (MFE). Data, mean ± SEM: *, p
    Figure Legend Snippet: Upregulation of CSC phenotypes in Akt-inhibitor-resistant tumor cells. ( A ) BT474 parental and Akt-inhibitor-resistant cells were seeded to plates in the absence of Akt inhibitor for 48 h. Cells were then treated with MK2206 or GDC0068 for 1 h. Whole-cell lysates were subjected to immunoblotting. * Resistant cells were cultured in the absence of Akt inhibitor for 3.5 weeks, before Western blot analysis. ( B ) Mammosphere forming assays of parental and resistant cells. BT474 cells were seeded for first-generation (1°) mammosphere formation assay. T47D cells were seeded for first generation of mammosphere formation assay, followed by second and then third generation (3°). Mammosphere forming efficiency (MFE). Data, mean ± SEM: *, p

    Techniques Used: Cell Culture, Western Blot, Tube Formation Assay

    Establishment and characterization of breast tumor cell lines resistant to Akt inhibitors. ( A ) Schematics of establishing Akt-inhibitor-resistant lines using high dose or step-wise method. ( B ) BT474 parental (black) and Akt-inhibitor-resistant (green) cells were seeded to 96-well plates and then treated with MK2206 or GDC0068 for 48 h. Cell viability was assessed by WST assays and calculated relative to the untreated cells. Data, mean ± standard error of the mean (SEM) with n = 3. ( C ) Parental and Akt-inhibitor-resistant cells were seeded to plates without Akt inhibitor for 48 h, followed by treatment with Akt inhibitor for 1 h. Whole-cell lysates were subjected to immunoblotting; p85 was used as a loading control.
    Figure Legend Snippet: Establishment and characterization of breast tumor cell lines resistant to Akt inhibitors. ( A ) Schematics of establishing Akt-inhibitor-resistant lines using high dose or step-wise method. ( B ) BT474 parental (black) and Akt-inhibitor-resistant (green) cells were seeded to 96-well plates and then treated with MK2206 or GDC0068 for 48 h. Cell viability was assessed by WST assays and calculated relative to the untreated cells. Data, mean ± standard error of the mean (SEM) with n = 3. ( C ) Parental and Akt-inhibitor-resistant cells were seeded to plates without Akt inhibitor for 48 h, followed by treatment with Akt inhibitor for 1 h. Whole-cell lysates were subjected to immunoblotting; p85 was used as a loading control.

    Techniques Used:

    4) Product Images from "β-Escin overcomes trastuzumab resistance in HER2-positive breast cancer by targeting cancer stem-like features"

    Article Title: β-Escin overcomes trastuzumab resistance in HER2-positive breast cancer by targeting cancer stem-like features

    Journal: Cancer Cell International

    doi: 10.1186/s12935-022-02713-9

    β-escin induces apoptosis in trastuzumab-sensitive and –resistant cells. A Chemical structure of β-escin. B The changes in morphology of BT474 and SKBR3 cells after treatment of β-escin (10-20 μM, 48 h) as observed by phase-contrast microscopy. C Representative phase-contrast images of JIMT-1 and MDA-MB-453 cells after treatment with β-escin (10-30 μM, 48 h). D , E Trastuzumab-sensitive SKBR3 and BT474 cells ( D ) and trastuzumab-resistant MDA-MB-453 and JIMT-1 cells ( E ) were treated with various concentrations of β-escin (1-100 μM) for 48 h, and cell viability was evaluated by MTS assay (**** p
    Figure Legend Snippet: β-escin induces apoptosis in trastuzumab-sensitive and –resistant cells. A Chemical structure of β-escin. B The changes in morphology of BT474 and SKBR3 cells after treatment of β-escin (10-20 μM, 48 h) as observed by phase-contrast microscopy. C Representative phase-contrast images of JIMT-1 and MDA-MB-453 cells after treatment with β-escin (10-30 μM, 48 h). D , E Trastuzumab-sensitive SKBR3 and BT474 cells ( D ) and trastuzumab-resistant MDA-MB-453 and JIMT-1 cells ( E ) were treated with various concentrations of β-escin (1-100 μM) for 48 h, and cell viability was evaluated by MTS assay (**** p

    Techniques Used: Microscopy, Multiple Displacement Amplification, MTS Assay

    β-escin-induced apoptosis is associated with activation of caspases and dysregulation of mitochondrial proteins. A , B The effects of β-escin (10-30 μM, 48 h) on apoptotic-related proteins. BT474 and JIMT-1 cells were treated with β-escin (10-20 μM and 20-30 μM, respectively) for 48 h. The expression of cleaved-caspase-7, cleaved-caspase-3, and cleaved-PARP were upregulated in both BT474 ( A ) and JIMT-1 cells ( B ). Quantitative graphs of protein content are shown in the right panels (* p
    Figure Legend Snippet: β-escin-induced apoptosis is associated with activation of caspases and dysregulation of mitochondrial proteins. A , B The effects of β-escin (10-30 μM, 48 h) on apoptotic-related proteins. BT474 and JIMT-1 cells were treated with β-escin (10-20 μM and 20-30 μM, respectively) for 48 h. The expression of cleaved-caspase-7, cleaved-caspase-3, and cleaved-PARP were upregulated in both BT474 ( A ) and JIMT-1 cells ( B ). Quantitative graphs of protein content are shown in the right panels (* p

    Techniques Used: Activation Assay, Expressing

    β-escin impairs CSC-like properties. A , B BT474 and JIMT-1 cells were treated with β-escin (10-30 μM) for 48 h and ALDH1 activity was assessed by flow cytometry. DEAB was used to define the baseline of Aldefluor-positive fluorescence. The quantitative graphs represent the percentage of Aldefluor-positive cells in BT474 (A, *** p
    Figure Legend Snippet: β-escin impairs CSC-like properties. A , B BT474 and JIMT-1 cells were treated with β-escin (10-30 μM) for 48 h and ALDH1 activity was assessed by flow cytometry. DEAB was used to define the baseline of Aldefluor-positive fluorescence. The quantitative graphs represent the percentage of Aldefluor-positive cells in BT474 (A, *** p

    Techniques Used: Activity Assay, Flow Cytometry, Fluorescence

    β-escin downregulates HER2, p95HER2, HER3 and Akt expression. A – D Western blots for HER2, p95HER2, phospho-HER2 (Y1221/1222), phospho-p95HER2, HER3, phospho-HER3 (Y1289), Akt, and phospho-Akt expression in BT474 ( A , C ) and JIMT-1 cells ( B , D ) following exposure to β-escin (10-20 μM and 20-30 μM, respectively) for 48 h. GAPDH was used as an internal loading control. The ratio of the protein content is represented in the right panels (* p
    Figure Legend Snippet: β-escin downregulates HER2, p95HER2, HER3 and Akt expression. A – D Western blots for HER2, p95HER2, phospho-HER2 (Y1221/1222), phospho-p95HER2, HER3, phospho-HER3 (Y1289), Akt, and phospho-Akt expression in BT474 ( A , C ) and JIMT-1 cells ( B , D ) following exposure to β-escin (10-20 μM and 20-30 μM, respectively) for 48 h. GAPDH was used as an internal loading control. The ratio of the protein content is represented in the right panels (* p

    Techniques Used: Expressing, Western Blot

    5) Product Images from "β-Escin overcomes trastuzumab resistance in HER2-positive breast cancer by targeting cancer stem-like features"

    Article Title: β-Escin overcomes trastuzumab resistance in HER2-positive breast cancer by targeting cancer stem-like features

    Journal: Cancer Cell International

    doi: 10.1186/s12935-022-02713-9

    β-escin induces apoptosis in trastuzumab-sensitive and –resistant cells. A Chemical structure of β-escin. B The changes in morphology of BT474 and SKBR3 cells after treatment of β-escin (10-20 μM, 48 h) as observed by phase-contrast microscopy. C Representative phase-contrast images of JIMT-1 and MDA-MB-453 cells after treatment with β-escin (10-30 μM, 48 h). D , E Trastuzumab-sensitive SKBR3 and BT474 cells ( D ) and trastuzumab-resistant MDA-MB-453 and JIMT-1 cells ( E ) were treated with various concentrations of β-escin (1-100 μM) for 48 h, and cell viability was evaluated by MTS assay (**** p
    Figure Legend Snippet: β-escin induces apoptosis in trastuzumab-sensitive and –resistant cells. A Chemical structure of β-escin. B The changes in morphology of BT474 and SKBR3 cells after treatment of β-escin (10-20 μM, 48 h) as observed by phase-contrast microscopy. C Representative phase-contrast images of JIMT-1 and MDA-MB-453 cells after treatment with β-escin (10-30 μM, 48 h). D , E Trastuzumab-sensitive SKBR3 and BT474 cells ( D ) and trastuzumab-resistant MDA-MB-453 and JIMT-1 cells ( E ) were treated with various concentrations of β-escin (1-100 μM) for 48 h, and cell viability was evaluated by MTS assay (**** p

    Techniques Used: Microscopy, Multiple Displacement Amplification, MTS Assay

    β-escin-induced apoptosis is associated with activation of caspases and dysregulation of mitochondrial proteins. A , B The effects of β-escin (10-30 μM, 48 h) on apoptotic-related proteins. BT474 and JIMT-1 cells were treated with β-escin (10-20 μM and 20-30 μM, respectively) for 48 h. The expression of cleaved-caspase-7, cleaved-caspase-3, and cleaved-PARP were upregulated in both BT474 ( A ) and JIMT-1 cells ( B ). Quantitative graphs of protein content are shown in the right panels (* p
    Figure Legend Snippet: β-escin-induced apoptosis is associated with activation of caspases and dysregulation of mitochondrial proteins. A , B The effects of β-escin (10-30 μM, 48 h) on apoptotic-related proteins. BT474 and JIMT-1 cells were treated with β-escin (10-20 μM and 20-30 μM, respectively) for 48 h. The expression of cleaved-caspase-7, cleaved-caspase-3, and cleaved-PARP were upregulated in both BT474 ( A ) and JIMT-1 cells ( B ). Quantitative graphs of protein content are shown in the right panels (* p

    Techniques Used: Activation Assay, Expressing

    β-escin impairs CSC-like properties. A , B BT474 and JIMT-1 cells were treated with β-escin (10-30 μM) for 48 h and ALDH1 activity was assessed by flow cytometry. DEAB was used to define the baseline of Aldefluor-positive fluorescence. The quantitative graphs represent the percentage of Aldefluor-positive cells in BT474 (A, *** p
    Figure Legend Snippet: β-escin impairs CSC-like properties. A , B BT474 and JIMT-1 cells were treated with β-escin (10-30 μM) for 48 h and ALDH1 activity was assessed by flow cytometry. DEAB was used to define the baseline of Aldefluor-positive fluorescence. The quantitative graphs represent the percentage of Aldefluor-positive cells in BT474 (A, *** p

    Techniques Used: Activity Assay, Flow Cytometry, Fluorescence

    β-escin downregulates HER2, p95HER2, HER3 and Akt expression. A – D Western blots for HER2, p95HER2, phospho-HER2 (Y1221/1222), phospho-p95HER2, HER3, phospho-HER3 (Y1289), Akt, and phospho-Akt expression in BT474 ( A , C ) and JIMT-1 cells ( B , D ) following exposure to β-escin (10-20 μM and 20-30 μM, respectively) for 48 h. GAPDH was used as an internal loading control. The ratio of the protein content is represented in the right panels (* p
    Figure Legend Snippet: β-escin downregulates HER2, p95HER2, HER3 and Akt expression. A – D Western blots for HER2, p95HER2, phospho-HER2 (Y1221/1222), phospho-p95HER2, HER3, phospho-HER3 (Y1289), Akt, and phospho-Akt expression in BT474 ( A , C ) and JIMT-1 cells ( B , D ) following exposure to β-escin (10-20 μM and 20-30 μM, respectively) for 48 h. GAPDH was used as an internal loading control. The ratio of the protein content is represented in the right panels (* p

    Techniques Used: Expressing, Western Blot

    6) Product Images from "β-Escin overcomes trastuzumab resistance in HER2-positive breast cancer by targeting cancer stem-like features"

    Article Title: β-Escin overcomes trastuzumab resistance in HER2-positive breast cancer by targeting cancer stem-like features

    Journal: Cancer Cell International

    doi: 10.1186/s12935-022-02713-9

    β-escin induces apoptosis in trastuzumab-sensitive and –resistant cells. A Chemical structure of β-escin. B The changes in morphology of BT474 and SKBR3 cells after treatment of β-escin (10-20 μM, 48 h) as observed by phase-contrast microscopy. C Representative phase-contrast images of JIMT-1 and MDA-MB-453 cells after treatment with β-escin (10-30 μM, 48 h). D , E Trastuzumab-sensitive SKBR3 and BT474 cells ( D ) and trastuzumab-resistant MDA-MB-453 and JIMT-1 cells ( E ) were treated with various concentrations of β-escin (1-100 μM) for 48 h, and cell viability was evaluated by MTS assay (**** p
    Figure Legend Snippet: β-escin induces apoptosis in trastuzumab-sensitive and –resistant cells. A Chemical structure of β-escin. B The changes in morphology of BT474 and SKBR3 cells after treatment of β-escin (10-20 μM, 48 h) as observed by phase-contrast microscopy. C Representative phase-contrast images of JIMT-1 and MDA-MB-453 cells after treatment with β-escin (10-30 μM, 48 h). D , E Trastuzumab-sensitive SKBR3 and BT474 cells ( D ) and trastuzumab-resistant MDA-MB-453 and JIMT-1 cells ( E ) were treated with various concentrations of β-escin (1-100 μM) for 48 h, and cell viability was evaluated by MTS assay (**** p

    Techniques Used: Microscopy, Multiple Displacement Amplification, MTS Assay

    β-escin-induced apoptosis is associated with activation of caspases and dysregulation of mitochondrial proteins. A , B The effects of β-escin (10-30 μM, 48 h) on apoptotic-related proteins. BT474 and JIMT-1 cells were treated with β-escin (10-20 μM and 20-30 μM, respectively) for 48 h. The expression of cleaved-caspase-7, cleaved-caspase-3, and cleaved-PARP were upregulated in both BT474 ( A ) and JIMT-1 cells ( B ). Quantitative graphs of protein content are shown in the right panels (* p
    Figure Legend Snippet: β-escin-induced apoptosis is associated with activation of caspases and dysregulation of mitochondrial proteins. A , B The effects of β-escin (10-30 μM, 48 h) on apoptotic-related proteins. BT474 and JIMT-1 cells were treated with β-escin (10-20 μM and 20-30 μM, respectively) for 48 h. The expression of cleaved-caspase-7, cleaved-caspase-3, and cleaved-PARP were upregulated in both BT474 ( A ) and JIMT-1 cells ( B ). Quantitative graphs of protein content are shown in the right panels (* p

    Techniques Used: Activation Assay, Expressing

    β-escin impairs CSC-like properties. A , B BT474 and JIMT-1 cells were treated with β-escin (10-30 μM) for 48 h and ALDH1 activity was assessed by flow cytometry. DEAB was used to define the baseline of Aldefluor-positive fluorescence. The quantitative graphs represent the percentage of Aldefluor-positive cells in BT474 (A, *** p
    Figure Legend Snippet: β-escin impairs CSC-like properties. A , B BT474 and JIMT-1 cells were treated with β-escin (10-30 μM) for 48 h and ALDH1 activity was assessed by flow cytometry. DEAB was used to define the baseline of Aldefluor-positive fluorescence. The quantitative graphs represent the percentage of Aldefluor-positive cells in BT474 (A, *** p

    Techniques Used: Activity Assay, Flow Cytometry, Fluorescence

    β-escin downregulates HER2, p95HER2, HER3 and Akt expression. A – D Western blots for HER2, p95HER2, phospho-HER2 (Y1221/1222), phospho-p95HER2, HER3, phospho-HER3 (Y1289), Akt, and phospho-Akt expression in BT474 ( A , C ) and JIMT-1 cells ( B , D ) following exposure to β-escin (10-20 μM and 20-30 μM, respectively) for 48 h. GAPDH was used as an internal loading control. The ratio of the protein content is represented in the right panels (* p
    Figure Legend Snippet: β-escin downregulates HER2, p95HER2, HER3 and Akt expression. A – D Western blots for HER2, p95HER2, phospho-HER2 (Y1221/1222), phospho-p95HER2, HER3, phospho-HER3 (Y1289), Akt, and phospho-Akt expression in BT474 ( A , C ) and JIMT-1 cells ( B , D ) following exposure to β-escin (10-20 μM and 20-30 μM, respectively) for 48 h. GAPDH was used as an internal loading control. The ratio of the protein content is represented in the right panels (* p

    Techniques Used: Expressing, Western Blot

    7) Product Images from "Impairment of IgG Fc functions promotes tumor progression and suppresses NK cell antitumor actions"

    Article Title: Impairment of IgG Fc functions promotes tumor progression and suppresses NK cell antitumor actions

    Journal: Communications Biology

    doi: 10.1038/s42003-022-03931-7

    Tumors containing scIgGs had an overall reduced immune cytokine levels. a Heat map of cytokine /chemokines levels detected in tumor lysates of PyMT+/scIgG+ vs PyMT+, n = 4. Tumor lysates were used for profiling a panel of 62 cytokines and chemokines (RayBio, c3 series, 62 cytokines) and tumor lysates used in each array were normalized with protein concentrations. b Heat map of cytokines/chemokines detected in tumor lysates of BT474-IdeS vs BT474. n = 6. Tumors with scIgGs had reduced cytokines more than 40% of the 62 panel RPPA (red color at X-axis) when compared to that in the control tumors. Fold of change (Y-axis) indicates ratio of cytokine levels in ( c ) PyMT+/scIgG+/PyMT+ and in ( d ) BT474-IdeS/ BT474. e , f Tumors with scIgGs had significantly lower IFN-γ levels in comparison with the control lysates. The levels of IFN-γ were analyzed using RPPA array data for ( e ) PyMT+/scIgG+ vs PyMT+ pair ( n = 4, p
    Figure Legend Snippet: Tumors containing scIgGs had an overall reduced immune cytokine levels. a Heat map of cytokine /chemokines levels detected in tumor lysates of PyMT+/scIgG+ vs PyMT+, n = 4. Tumor lysates were used for profiling a panel of 62 cytokines and chemokines (RayBio, c3 series, 62 cytokines) and tumor lysates used in each array were normalized with protein concentrations. b Heat map of cytokines/chemokines detected in tumor lysates of BT474-IdeS vs BT474. n = 6. Tumors with scIgGs had reduced cytokines more than 40% of the 62 panel RPPA (red color at X-axis) when compared to that in the control tumors. Fold of change (Y-axis) indicates ratio of cytokine levels in ( c ) PyMT+/scIgG+/PyMT+ and in ( d ) BT474-IdeS/ BT474. e , f Tumors with scIgGs had significantly lower IFN-γ levels in comparison with the control lysates. The levels of IFN-γ were analyzed using RPPA array data for ( e ) PyMT+/scIgG+ vs PyMT+ pair ( n = 4, p

    Techniques Used:

    Tumors grown in FcγR KO mice mirrored NK immune suppression shown in scIgG containing tumors. CD16 binding signals in high scIgG containing tumor lysates (OD450 nm/μg protein) were significantly lower than that in tumor lysates. Graph ( a ) shows comparison between BT474-IdeS and BT474 control tumor model and ( b ) shows comparison of the CD16 bindings between 4T1-IdeS vs 4T1 tumors. CD16 bindings were determined using ELISA and the binding intensities were calculated by (OD450 nm/μg protein), n = 6, p
    Figure Legend Snippet: Tumors grown in FcγR KO mice mirrored NK immune suppression shown in scIgG containing tumors. CD16 binding signals in high scIgG containing tumor lysates (OD450 nm/μg protein) were significantly lower than that in tumor lysates. Graph ( a ) shows comparison between BT474-IdeS and BT474 control tumor model and ( b ) shows comparison of the CD16 bindings between 4T1-IdeS vs 4T1 tumors. CD16 bindings were determined using ELISA and the binding intensities were calculated by (OD450 nm/μg protein), n = 6, p

    Techniques Used: Mouse Assay, Binding Assay, Enzyme-linked Immunosorbent Assay

    AHA rescued NK cell infiltration to tumors and showed antitumor activity in scIgG containing tumor models. a AHA treatment significantly inhibited tumor growth in mouse xenograft tumor model of BT474-IdeS xenograft tumor model. Tumor growth curves of each individual mice are shown and blue arrow indicates starting antibody treatment, n = 5, *** p
    Figure Legend Snippet: AHA rescued NK cell infiltration to tumors and showed antitumor activity in scIgG containing tumor models. a AHA treatment significantly inhibited tumor growth in mouse xenograft tumor model of BT474-IdeS xenograft tumor model. Tumor growth curves of each individual mice are shown and blue arrow indicates starting antibody treatment, n = 5, *** p

    Techniques Used: Activity Assay, Mouse Assay

    Elevated levels of tumor scIgGs promoted tumor growth and tumor initiation. a – c Detection of scIgGs in tumor tissues using immune florescence (IF) imaging method. A representative image is shown for each tumor group and scale bars are indicated on each images. Five tumor slides ( n = 5) in each group were stained using IF imaging method and 4 images were collected per tumor slides. Images in ( a ) shows scIgG staining between BT474-IdeS and control BT474 tumors. Images in ( b ) shows scIgG staining between 4T1-IdeS and control 4T1 tumors. Images in ( c ) shows comparison of scIgG staining between PyMT+/scIgG+ tumors and PyMT+ control tumors. AHA monoclonal antibody (2095-2) was conjugated with DyLight (red color) using a commercial conjugation kit (Thermo Fisher) for detection of scIgGs in tumor tissues. d Tumor growth of BT474-IdeS cancer cells in vivo was significantly faster than the control (BT474) tumors, p
    Figure Legend Snippet: Elevated levels of tumor scIgGs promoted tumor growth and tumor initiation. a – c Detection of scIgGs in tumor tissues using immune florescence (IF) imaging method. A representative image is shown for each tumor group and scale bars are indicated on each images. Five tumor slides ( n = 5) in each group were stained using IF imaging method and 4 images were collected per tumor slides. Images in ( a ) shows scIgG staining between BT474-IdeS and control BT474 tumors. Images in ( b ) shows scIgG staining between 4T1-IdeS and control 4T1 tumors. Images in ( c ) shows comparison of scIgG staining between PyMT+/scIgG+ tumors and PyMT+ control tumors. AHA monoclonal antibody (2095-2) was conjugated with DyLight (red color) using a commercial conjugation kit (Thermo Fisher) for detection of scIgGs in tumor tissues. d Tumor growth of BT474-IdeS cancer cells in vivo was significantly faster than the control (BT474) tumors, p

    Techniques Used: Imaging, Staining, Conjugation Assay, In Vivo

    8) Product Images from "Analysis of RANK-c interaction partners identifies TRAF3 as a critical regulator of breast cancer aggressiveness"

    Article Title: Analysis of RANK-c interaction partners identifies TRAF3 as a critical regulator of breast cancer aggressiveness

    Journal: Neoplasia (New York, N.Y.)

    doi: 10.1016/j.neo.2022.100836

    Forced expression of TRAF3 in BT474-RANK-c cell is able to reverse the aggressive cell phenotype. (A) Colony formation assay and quantification of BT474-RANK-c and parental cells (control) transiently transfected with a TRAF3 plasmid or an inactive TRAF3 mutant plasmid (TRAF3mut). (B) Transwell migration assay and quantification of BT474-RANK-c and control cells after transient transfection with the indicated plasmid constructs for TRAF3 (TRAF3 and TRAF3mut). (C) Matrigel invasion assay and quantification of BT474-RANK-c and control cells after transient transfection with the indicated plasmid constructs (as for migration).
    Figure Legend Snippet: Forced expression of TRAF3 in BT474-RANK-c cell is able to reverse the aggressive cell phenotype. (A) Colony formation assay and quantification of BT474-RANK-c and parental cells (control) transiently transfected with a TRAF3 plasmid or an inactive TRAF3 mutant plasmid (TRAF3mut). (B) Transwell migration assay and quantification of BT474-RANK-c and control cells after transient transfection with the indicated plasmid constructs for TRAF3 (TRAF3 and TRAF3mut). (C) Matrigel invasion assay and quantification of BT474-RANK-c and control cells after transient transfection with the indicated plasmid constructs (as for migration).

    Techniques Used: Expressing, Colony Assay, Transfection, Plasmid Preparation, Mutagenesis, Transwell Migration Assay, Construct, Invasion Assay, Migration

    RANK-c differentially affects multiple signalling pathways in SKBR3 and BT474 cells. (A) Western blot analyses for the indicated proteins in SKBR3 and BT474 cells (RANK-c and control). (B) Representative images and quantification of immunofluorescence staining of SKBR3-RANK-c, BT474-RANK-c and control cells for RANK-c and p65 protein localization. Split channels at 40x magnification. (C) Western blot analysis of RANK-c expressing SKBR3 and BT474 cell lysates immunoprecipitated with an antibody against RANK (AF683) and blotted for endogenous TRAF2 protein.
    Figure Legend Snippet: RANK-c differentially affects multiple signalling pathways in SKBR3 and BT474 cells. (A) Western blot analyses for the indicated proteins in SKBR3 and BT474 cells (RANK-c and control). (B) Representative images and quantification of immunofluorescence staining of SKBR3-RANK-c, BT474-RANK-c and control cells for RANK-c and p65 protein localization. Split channels at 40x magnification. (C) Western blot analysis of RANK-c expressing SKBR3 and BT474 cell lysates immunoprecipitated with an antibody against RANK (AF683) and blotted for endogenous TRAF2 protein.

    Techniques Used: Western Blot, Immunofluorescence, Staining, Expressing, Immunoprecipitation

    RANK-c expression in SKBR3 and BT474 cells differentially affects aggressive properties. (A) Phase-contrast images of 2D cell cultures in standard media depicting morphological changes upon RANK-c expression. SKBR3-RANK-c cells lose their fibroblast-like appearance, while in the contrary BT474 cells lose coherence and attain a spindle-cell morphology. (B) Western blot of SKBR3 and BT474 cells extracts showing the downregulation and upregulation of Fibronectin and Snail upon RANK-c expression, respectively. (C) Representative images and quantification of colony formation by SKBR3-RANK-c and BT474-RANK-c cells in relevance to the respective control cells ( n = 3 wells per group). (D) Transwell migration quantification for SKBR3-RANK-c, BT474-RANK-c and control cells ( n = = 3). (E) Matrigel invasion assay and quantification for SKBR3-RANK-c, BT474-RANK-c and control cells ( n = = 3).
    Figure Legend Snippet: RANK-c expression in SKBR3 and BT474 cells differentially affects aggressive properties. (A) Phase-contrast images of 2D cell cultures in standard media depicting morphological changes upon RANK-c expression. SKBR3-RANK-c cells lose their fibroblast-like appearance, while in the contrary BT474 cells lose coherence and attain a spindle-cell morphology. (B) Western blot of SKBR3 and BT474 cells extracts showing the downregulation and upregulation of Fibronectin and Snail upon RANK-c expression, respectively. (C) Representative images and quantification of colony formation by SKBR3-RANK-c and BT474-RANK-c cells in relevance to the respective control cells ( n = 3 wells per group). (D) Transwell migration quantification for SKBR3-RANK-c, BT474-RANK-c and control cells ( n = = 3). (E) Matrigel invasion assay and quantification for SKBR3-RANK-c, BT474-RANK-c and control cells ( n = = 3).

    Techniques Used: Expressing, Western Blot, Migration, Invasion Assay

    TRAF3 is absent from the RANK-c/TRAF2 complex in BT474 cells. (A) Volcano plots of RANK-c-enriched samples compared to their controls in SKBR3 and BT474 cells. The lines represent significance cut-offs of a 1% and 5% FDR with an S0 value of 0.1. All proteins identified in this experiment are listed in Supplementary Table 1. (B) Western blot analysis of SKBR3-RANK-c and BT474-RANK-c cell lysates immunoprecipitated with an antibody against RANK (AF683) and blotted for endogenous TRAF2 and TRAF3 proteins. (C) Western blot analysis of 293T cells transiently transfected with the indicated plasmid constructs and subsequently lysed and immunoprecipitated with an antibody against TRAF2 (sc-136999) and blotted for RANK-c and TRAF3 proteins.
    Figure Legend Snippet: TRAF3 is absent from the RANK-c/TRAF2 complex in BT474 cells. (A) Volcano plots of RANK-c-enriched samples compared to their controls in SKBR3 and BT474 cells. The lines represent significance cut-offs of a 1% and 5% FDR with an S0 value of 0.1. All proteins identified in this experiment are listed in Supplementary Table 1. (B) Western blot analysis of SKBR3-RANK-c and BT474-RANK-c cell lysates immunoprecipitated with an antibody against RANK (AF683) and blotted for endogenous TRAF2 and TRAF3 proteins. (C) Western blot analysis of 293T cells transiently transfected with the indicated plasmid constructs and subsequently lysed and immunoprecipitated with an antibody against TRAF2 (sc-136999) and blotted for RANK-c and TRAF3 proteins.

    Techniques Used: Western Blot, Immunoprecipitation, Transfection, Plasmid Preparation, Construct

    9) Product Images from "A comprehensive role evaluation and mechanism exploration of POGLUT2 in pan-cancer"

    Article Title: A comprehensive role evaluation and mechanism exploration of POGLUT2 in pan-cancer

    Journal: Frontiers in Oncology

    doi: 10.3389/fonc.2022.962540

    Gene and protein expression evaluation of POGLUT2 in breast cancer cells and tissues. (A) Gene expression of POGLUT2 was determined in four breast cancer cell lines, including MDA-MB-468, MDA-MB-231, MCF-7, and BT474, and highest expression was found in MCF-7 cells. (B) Protein expression of POGLUT2 was determined in four breast cancer cell lines, including MDA-MB-468, MDA-MB-231, MCF-7, and BT474, and highest expression was found in MDA-MB-231 cells. (C) Immunohistochemistry analysis of POGLUT2 in breast cancer tissues compared to control tissues. (D) Significant POGLUT2 expression downregulation was found in MCF-7 cells after siRNA transfection. (E) Significantly POGLUT2 expression downregulation was found in MDA-MB-231 cells after siRNA transfection. (F) Significantly POGLUT2 protein level downregulation was found in MCF-7 cells after siRNA transfection. (G) Significantly POGLUT2 protein level downregulation was found in MDA-MB-231 cells after siRNA transfection. * p
    Figure Legend Snippet: Gene and protein expression evaluation of POGLUT2 in breast cancer cells and tissues. (A) Gene expression of POGLUT2 was determined in four breast cancer cell lines, including MDA-MB-468, MDA-MB-231, MCF-7, and BT474, and highest expression was found in MCF-7 cells. (B) Protein expression of POGLUT2 was determined in four breast cancer cell lines, including MDA-MB-468, MDA-MB-231, MCF-7, and BT474, and highest expression was found in MDA-MB-231 cells. (C) Immunohistochemistry analysis of POGLUT2 in breast cancer tissues compared to control tissues. (D) Significant POGLUT2 expression downregulation was found in MCF-7 cells after siRNA transfection. (E) Significantly POGLUT2 expression downregulation was found in MDA-MB-231 cells after siRNA transfection. (F) Significantly POGLUT2 protein level downregulation was found in MCF-7 cells after siRNA transfection. (G) Significantly POGLUT2 protein level downregulation was found in MDA-MB-231 cells after siRNA transfection. * p

    Techniques Used: Expressing, Multiple Displacement Amplification, Immunohistochemistry, Transfection

    10) Product Images from "Quantifying the Effects of Combination Trastuzumab and Radiation Therapy in Human Epidermal Growth Factor Receptor 2-Positive Breast Cancer"

    Article Title: Quantifying the Effects of Combination Trastuzumab and Radiation Therapy in Human Epidermal Growth Factor Receptor 2-Positive Breast Cancer

    Journal: Cancers

    doi: 10.3390/cancers14174234

    Differences in BT474 cell growth with trastuzumab administered before, at the same time, or after radiation therapy. Treatment with trastuzumab before radiation did not significantly alter cell response to therapy compared with trastuzumab treatment at the same time ( p = 1.00) or treatment after radiation ( p = 0.98). Treatment with trastuzumab at the same time did not significantly alter the cell response to therapy compared with treatment after radiation ( p = 0.93). Altering the order of dosing trastuzumab and radiation did not alter the cell response to treatment in vitro.
    Figure Legend Snippet: Differences in BT474 cell growth with trastuzumab administered before, at the same time, or after radiation therapy. Treatment with trastuzumab before radiation did not significantly alter cell response to therapy compared with trastuzumab treatment at the same time ( p = 1.00) or treatment after radiation ( p = 0.98). Treatment with trastuzumab at the same time did not significantly alter the cell response to therapy compared with treatment after radiation ( p = 0.93). Altering the order of dosing trastuzumab and radiation did not alter the cell response to treatment in vitro.

    Techniques Used: In Vitro

    Interaction index calculated per time point over one week of treatment. All groups were treated with 0.01 ng/mL of trastuzumab. Graphs display the interaction indexes of ( A ) BT474 cells with 5 Gy radiation and trastuzumab; ( B ) BT474 cells with 10 Gy radiation and trastuzumab; ( C ) SKBR3 cells with 5 Gy radiation and trastuzumab; ( D ) SKBR3 cells with 10 Gy radiation and trastuzumab; ( E ) MDA-MB-231 cells with 5 Gy radiation and trastuzumab; and ( F ) MDA-MB-231 cells with 10 Gy radiation and trastuzumab. No group had an interaction index significantly above or below 0 at any time point, indicating additive treatment effects.
    Figure Legend Snippet: Interaction index calculated per time point over one week of treatment. All groups were treated with 0.01 ng/mL of trastuzumab. Graphs display the interaction indexes of ( A ) BT474 cells with 5 Gy radiation and trastuzumab; ( B ) BT474 cells with 10 Gy radiation and trastuzumab; ( C ) SKBR3 cells with 5 Gy radiation and trastuzumab; ( D ) SKBR3 cells with 10 Gy radiation and trastuzumab; ( E ) MDA-MB-231 cells with 5 Gy radiation and trastuzumab; and ( F ) MDA-MB-231 cells with 10 Gy radiation and trastuzumab. No group had an interaction index significantly above or below 0 at any time point, indicating additive treatment effects.

    Techniques Used: Multiple Displacement Amplification

    HER2 quantification in BT474, SKBR3, and MDA-MB-231 cells. ( A ) Expression of HER2 protein (185 kDa) in each cell line. β-actin (42 kDa) was used as an internal control. Visible protein bands are seen in the BT474 and SKBR3 HER2+ cell lines, while MDA-MB-231 shows little to no expression. ( B ) HER2:β-actin ratio in each cell line. BT474 and SKBR3 cells have ratios of 1.41 and 1.46, respectively. MDA-MB-231 has a ratio of 0.08, confirming no HER2 amplification. The uncropped blots are shown in Supplementary Figure S2 .
    Figure Legend Snippet: HER2 quantification in BT474, SKBR3, and MDA-MB-231 cells. ( A ) Expression of HER2 protein (185 kDa) in each cell line. β-actin (42 kDa) was used as an internal control. Visible protein bands are seen in the BT474 and SKBR3 HER2+ cell lines, while MDA-MB-231 shows little to no expression. ( B ) HER2:β-actin ratio in each cell line. BT474 and SKBR3 cells have ratios of 1.41 and 1.46, respectively. MDA-MB-231 has a ratio of 0.08, confirming no HER2 amplification. The uncropped blots are shown in Supplementary Figure S2 .

    Techniques Used: Multiple Displacement Amplification, Expressing, Amplification

    Cell proliferation over one week after radiation, trastuzumab, or combination treatment. All trastuzumab single-agent and combination groups were treated with 0.1 ng/mL of trastuzumab from 0–48 h. All radiation single-agent and combination groups were treated with 5 or 10 Gy radiation on Day 0. Graphs display the proliferation of: ( A ) BT474 cells after trastuzumab, 5 Gy radiation, and combination treatment, ( B ) BT474 cells after trastuzumab, 10 Gy radiation, and combination treatment, ( C ) SKBR3 cells after trastuzumab, 5 Gy radiation, and combination treatment, ( D ) SKBR3 cells after trastuzumab, 10 Gy radiation, and combination treatment, ( E ) MDA-MB-231 cells after trastuzumab, 5 Gy radiation, and combination treatment, and ( F ) MDA-MB-231 cells after trastuzumab, 10 Gy radiation, and combination treatment. No significant difference was observed in the control MDA-MB-231 cell proliferation and cells treated with single-agent trastuzumab ( p = 0.88, ( E , F )). No significant difference in cell proliferation was observed between cells treated with single-agent radiation and cells treated with combination treatment ( p = 0.84, ( E ) and p = 0.80, ( F )).
    Figure Legend Snippet: Cell proliferation over one week after radiation, trastuzumab, or combination treatment. All trastuzumab single-agent and combination groups were treated with 0.1 ng/mL of trastuzumab from 0–48 h. All radiation single-agent and combination groups were treated with 5 or 10 Gy radiation on Day 0. Graphs display the proliferation of: ( A ) BT474 cells after trastuzumab, 5 Gy radiation, and combination treatment, ( B ) BT474 cells after trastuzumab, 10 Gy radiation, and combination treatment, ( C ) SKBR3 cells after trastuzumab, 5 Gy radiation, and combination treatment, ( D ) SKBR3 cells after trastuzumab, 10 Gy radiation, and combination treatment, ( E ) MDA-MB-231 cells after trastuzumab, 5 Gy radiation, and combination treatment, and ( F ) MDA-MB-231 cells after trastuzumab, 10 Gy radiation, and combination treatment. No significant difference was observed in the control MDA-MB-231 cell proliferation and cells treated with single-agent trastuzumab ( p = 0.88, ( E , F )). No significant difference in cell proliferation was observed between cells treated with single-agent radiation and cells treated with combination treatment ( p = 0.84, ( E ) and p = 0.80, ( F )).

    Techniques Used: Multiple Displacement Amplification

    11) Product Images from "Extracellular Vesicles Carrying miR-887-3p Promote Breast Cancer Cell Drug Resistance by Targeting BTBD7 and Activating the Notch1/Hes1 Signaling Pathway"

    Article Title: Extracellular Vesicles Carrying miR-887-3p Promote Breast Cancer Cell Drug Resistance by Targeting BTBD7 and Activating the Notch1/Hes1 Signaling Pathway

    Journal: Disease Markers

    doi: 10.1155/2022/5762686

    miR-887-3p targeted BTBD7. (a) Venn map of the intersection of miR-887-3p downstream target genes predicted by RNAInter, Targetscan, and ENCORI databases. (b) Coexpression network of candidate genes. (c) miR-887-3p targeting site in BTBD7 3′UTR predicted by Starbase. (d) Luciferase reporter plasmid containing BTBD7-wt or BTBD7-mut was transfected into 293T cells together with miR-887-3p in parallel with an miR-NC plasmid vector. (e) Enrichment of miR-887-3p on the BTBD7 was detected by RNA pull down assay, relative to antisense-oligos. (f) ECORI Pan-Cer assessed the BTBD7 expression in normal tissues and primary tumors. Relative BTBD7 mRNA expression (g) and protein level (h) in MCF-7, BT474, and HCC1937 cells were determined by RT-qPCR and western blot analysis under MDA-MB-231-derived EV treatment. The miR-887-3p was normalized to U6 while the BTBD7 was normalized to GAPDH. Data are expressed as the mean ± standard deviation. One-way ANOVA and Tukey's multiple comparison test were used. ∗ p
    Figure Legend Snippet: miR-887-3p targeted BTBD7. (a) Venn map of the intersection of miR-887-3p downstream target genes predicted by RNAInter, Targetscan, and ENCORI databases. (b) Coexpression network of candidate genes. (c) miR-887-3p targeting site in BTBD7 3′UTR predicted by Starbase. (d) Luciferase reporter plasmid containing BTBD7-wt or BTBD7-mut was transfected into 293T cells together with miR-887-3p in parallel with an miR-NC plasmid vector. (e) Enrichment of miR-887-3p on the BTBD7 was detected by RNA pull down assay, relative to antisense-oligos. (f) ECORI Pan-Cer assessed the BTBD7 expression in normal tissues and primary tumors. Relative BTBD7 mRNA expression (g) and protein level (h) in MCF-7, BT474, and HCC1937 cells were determined by RT-qPCR and western blot analysis under MDA-MB-231-derived EV treatment. The miR-887-3p was normalized to U6 while the BTBD7 was normalized to GAPDH. Data are expressed as the mean ± standard deviation. One-way ANOVA and Tukey's multiple comparison test were used. ∗ p

    Techniques Used: Luciferase, Plasmid Preparation, Transfection, Pull Down Assay, Expressing, Quantitative RT-PCR, Western Blot, Multiple Displacement Amplification, Derivative Assay, Standard Deviation

    MDA-MB-231-derived EVs promoted BC cell drug resistance. (a) The survival of MCF-7, HCC1937, and BT474 cells in different concentrations of doxorubicin (Dox), cisplatin (Cis), and fulvestrant (Ful) was detected using CCK-8 assay. Then, the BC cells were treated with 15 μ g/mL MDA-MB-231-derived EVs (EV-group) or MDA-MB-231 supernatant supplemented with GW4869 (NC group), or PBS as a negative control (blank group). (c) Fluorescence microscope showed the uptake of EVs (15 μ g/mL) labeled with the red fluorescent dye PKH26 by MCF-7, BT474, and HCC1937 cells after 24 h of stimulation. Next, MDA-MB-231-derived EV-treated parental or drug-resistant BC cells were exposed to 0.5 μ M doxorubicin, 2.5 μ M cisplatin, and 0.5 μ M fulvestrant, respectively. Then, CCK-8 assay (c), colony formation assay (d), and flow cytometry (e) were performed to determine the effect of MDA-MB-231-derived EVs on BC cell drug resistance. Data are expressed as the mean ± standard deviation; one-way ANOVA and Tukey's multiple comparison test were used to determine statistical significance. ∗ p
    Figure Legend Snippet: MDA-MB-231-derived EVs promoted BC cell drug resistance. (a) The survival of MCF-7, HCC1937, and BT474 cells in different concentrations of doxorubicin (Dox), cisplatin (Cis), and fulvestrant (Ful) was detected using CCK-8 assay. Then, the BC cells were treated with 15 μ g/mL MDA-MB-231-derived EVs (EV-group) or MDA-MB-231 supernatant supplemented with GW4869 (NC group), or PBS as a negative control (blank group). (c) Fluorescence microscope showed the uptake of EVs (15 μ g/mL) labeled with the red fluorescent dye PKH26 by MCF-7, BT474, and HCC1937 cells after 24 h of stimulation. Next, MDA-MB-231-derived EV-treated parental or drug-resistant BC cells were exposed to 0.5 μ M doxorubicin, 2.5 μ M cisplatin, and 0.5 μ M fulvestrant, respectively. Then, CCK-8 assay (c), colony formation assay (d), and flow cytometry (e) were performed to determine the effect of MDA-MB-231-derived EVs on BC cell drug resistance. Data are expressed as the mean ± standard deviation; one-way ANOVA and Tukey's multiple comparison test were used to determine statistical significance. ∗ p

    Techniques Used: Multiple Displacement Amplification, Derivative Assay, CCK-8 Assay, Negative Control, Fluorescence, Microscopy, Labeling, Colony Assay, Flow Cytometry, Standard Deviation

    MDA-MB-231-derived EVs carried miR-887-3p into BC cells. ECORI Pan-Cer database analyzed the expression (a) and prognosis (b) of miR-887-3p in BC. EVs were treated with RNase (RNase-group) and RNase+SDS (RNase+SDS-group), respectively. miR-887-3p expression was detected using RT-qPCR (c). Next, miR-Inhibitor was transfected into MDA-MB-231 cells while Mock miR served as a negative control. Then, EVs (EV-NC group and EV-inhi group) were extracted as described in a method. RT-qPCR was performed to determine miR-887-3p expression in MDA-MB-231 cells (d) and EVs (e). MCF-7, BT474, and HCC1937 cells were treated with EVs in different groups, and then RT-qPCR was performed to determine miR-887-3p expression (f). Data were expressed as the mean ± standard deviation. In (f), two-way ANOVA and Tukey's multiple comparison test were used to determine statistical significance, whereas in (c)–(e) one-way ANOVA and Tukey's multiple comparison test were used. ∗ p
    Figure Legend Snippet: MDA-MB-231-derived EVs carried miR-887-3p into BC cells. ECORI Pan-Cer database analyzed the expression (a) and prognosis (b) of miR-887-3p in BC. EVs were treated with RNase (RNase-group) and RNase+SDS (RNase+SDS-group), respectively. miR-887-3p expression was detected using RT-qPCR (c). Next, miR-Inhibitor was transfected into MDA-MB-231 cells while Mock miR served as a negative control. Then, EVs (EV-NC group and EV-inhi group) were extracted as described in a method. RT-qPCR was performed to determine miR-887-3p expression in MDA-MB-231 cells (d) and EVs (e). MCF-7, BT474, and HCC1937 cells were treated with EVs in different groups, and then RT-qPCR was performed to determine miR-887-3p expression (f). Data were expressed as the mean ± standard deviation. In (f), two-way ANOVA and Tukey's multiple comparison test were used to determine statistical significance, whereas in (c)–(e) one-way ANOVA and Tukey's multiple comparison test were used. ∗ p

    Techniques Used: Multiple Displacement Amplification, Derivative Assay, Expressing, Quantitative RT-PCR, Transfection, Negative Control, Standard Deviation

    MDA-MB-231-derived EVs carrying miR-887-3p targeted BTBD7 and activated Notch1/Hes1 signaling pathway to promote BC cell drug resistance. MCF-7, HCC1937, and BT474 cells were treated with EVs and transfected with oe-BTBD7. (a) RT-qPCR was performed to determine BTBD7 mRNA expression. CCK-8 assay (b), colony formation assay (c), and flow cytometry (d) were performed to determine the effects of BTBD7 on BC cell drug resistance. Western blot analysis (e) was performed to determine Notch1 and Hes1 protein level in MCF-7, BT474, and HCC1937 cells under MDA-MB-231-derived EV treatments. Data are expressed as the mean ± standard deviation. One-way ANOVA and Tukey's multiple comparison test were used. ∗ p
    Figure Legend Snippet: MDA-MB-231-derived EVs carrying miR-887-3p targeted BTBD7 and activated Notch1/Hes1 signaling pathway to promote BC cell drug resistance. MCF-7, HCC1937, and BT474 cells were treated with EVs and transfected with oe-BTBD7. (a) RT-qPCR was performed to determine BTBD7 mRNA expression. CCK-8 assay (b), colony formation assay (c), and flow cytometry (d) were performed to determine the effects of BTBD7 on BC cell drug resistance. Western blot analysis (e) was performed to determine Notch1 and Hes1 protein level in MCF-7, BT474, and HCC1937 cells under MDA-MB-231-derived EV treatments. Data are expressed as the mean ± standard deviation. One-way ANOVA and Tukey's multiple comparison test were used. ∗ p

    Techniques Used: Multiple Displacement Amplification, Derivative Assay, Transfection, Quantitative RT-PCR, Expressing, CCK-8 Assay, Colony Assay, Flow Cytometry, Western Blot, Standard Deviation

    12) Product Images from "Transcriptome profiling and proteomic validation reveals targets of the androgen receptor signaling in the BT-474 breast cancer cell line"

    Article Title: Transcriptome profiling and proteomic validation reveals targets of the androgen receptor signaling in the BT-474 breast cancer cell line

    Journal: Clinical Proteomics

    doi: 10.1186/s12014-022-09352-2

    Pearson correlation analysis between the transcript and protein levels of DHT-regulated genes in BT474 cells. The X axis shows the transcriptomic (log 2 RPKM ratio) and the Y axis the proteomic (log 2 PRM ratio) levels of DHT-treated BT474 cells. The graphs A and B show the comparison of the RNA sequencing results with the two PRM experiments, respectively. Pearson correlation was calculated using the GraphPad software
    Figure Legend Snippet: Pearson correlation analysis between the transcript and protein levels of DHT-regulated genes in BT474 cells. The X axis shows the transcriptomic (log 2 RPKM ratio) and the Y axis the proteomic (log 2 PRM ratio) levels of DHT-treated BT474 cells. The graphs A and B show the comparison of the RNA sequencing results with the two PRM experiments, respectively. Pearson correlation was calculated using the GraphPad software

    Techniques Used: RNA Sequencing Assay, Software

    Proposed action of AR in the AR + ER + BCa cell line, BT474. Treatment of BT474 cells with the androgen DHT caused upregulation of known and unknown A-regulated genes, including genes related to fatty acid metabolism and tumor-suppressor genes, such as ZBTB16 and CLDN8 . At the same time, AR suppressed directly or indirectly the expression of genes related to various pathways, such as WNT signaling, and ER-responsive pathways, resulting in decreased cell proliferation and tumor growth. Boxes in red and blue color represent upregulated and downregulated pathways, respectively
    Figure Legend Snippet: Proposed action of AR in the AR + ER + BCa cell line, BT474. Treatment of BT474 cells with the androgen DHT caused upregulation of known and unknown A-regulated genes, including genes related to fatty acid metabolism and tumor-suppressor genes, such as ZBTB16 and CLDN8 . At the same time, AR suppressed directly or indirectly the expression of genes related to various pathways, such as WNT signaling, and ER-responsive pathways, resulting in decreased cell proliferation and tumor growth. Boxes in red and blue color represent upregulated and downregulated pathways, respectively

    Techniques Used: Expressing

    Project pipeline. A Workflow of the transcriptomic analysis of hormonally treated BT474 cells. Cells were cultured and treated with hormones or ethanol (used as control) for 24 h. Cell pellets were collected and subjected to total RNA isolation. GSEA and Cytoscape software was used for the functional and pathway analysis of the differentially expressed genes. DHT dihydrotestosterone (10 nM), PROG progesterone (10 nM). B Principal Component Analysis (PCA) of BT474 RNA sequencing in three treatment conditions (Ethanol, DHT, and PROG). Each condition was performed in triplicates and groups clearly cluster. DHT dihydrotestosterone (10 nM), PROG progesterone (10 nM). C Experimental workflow of targeted PRM proteomics and data analysis for the validation of select androgen-regulated proteins in the BT474 cell line. For details see under “methods”
    Figure Legend Snippet: Project pipeline. A Workflow of the transcriptomic analysis of hormonally treated BT474 cells. Cells were cultured and treated with hormones or ethanol (used as control) for 24 h. Cell pellets were collected and subjected to total RNA isolation. GSEA and Cytoscape software was used for the functional and pathway analysis of the differentially expressed genes. DHT dihydrotestosterone (10 nM), PROG progesterone (10 nM). B Principal Component Analysis (PCA) of BT474 RNA sequencing in three treatment conditions (Ethanol, DHT, and PROG). Each condition was performed in triplicates and groups clearly cluster. DHT dihydrotestosterone (10 nM), PROG progesterone (10 nM). C Experimental workflow of targeted PRM proteomics and data analysis for the validation of select androgen-regulated proteins in the BT474 cell line. For details see under “methods”

    Techniques Used: Cell Culture, Isolation, Software, Functional Assay, RNA Sequencing Assay

    Hormonally regulated changes in the BT474 transcriptome. A Pie charts illustrating the different transcript types that are hormonally regulated in BT474 cells. In both conditions, DHT- and PROG-treated cells, 79% of the transcripts are protein-coding, 5% long non-coding RNAs (lncRNA), 2% microRNAs (miRNA), 2% small non-coding RNA (ncRNA), 2% pseudogenes, and the remaining 11% belong to other categories and gene ID duplications. *snoRNA, snRNA, scaRNA, scRNA, miscRNA. **Polymorphic_pseudogene, processed_pseudogene, transcribed_processed_pseudogene, transcribed_unitary_pseudogene, transcribed_unprocessed_ pseudogene, unitary_pseudogene, unprocessed_pseudogene. ***TEC (To be experimentally confirmed), ribosome The transcript categorization was performed on Ensembl website, using the bioinformatics tool, Biomart. B Histograms demonstrating the FC (fold change) distribution of DEGs in the transcript list, C Volcano plots illustrating the most significant differentially expressed genes. Red color shows the upregulated genes and blue color shows the downregulated genes. D Heatmap illustrating the differential expression profiles in DHT-treated and PROG-treated cells, respectively, compared to controls. Red color represents the upregulated genes, whereas the blue color represents the downregulated genes. Grey color corresponds to the genes that were not expressed, or the ratio could not be assessed in this experiment. All graphs were constructed in Perseus software using the log 2 FC values derived from the expression comparison between the hormone-treated and control cells. E , F Venn diagram representing the number and overlapping of DEGs in DHT-treated BT474 cells with known AR-regulated and cancer/BCa-related genes. The identification of the cancer/BCa related enriched genes were identified with the Gene Ontology annotation ( E ) and the KEGG analysis ( F ) of DAVID (2021 Update) online tool. G Comparison of gene set numbers identified in GSEA analysis of the RNA-seq experiment in DHT and PROG-treated BT474 cells. For the GSEA (v.3.0) analysis, only gene sets that contained 15–200 genes were used (q-value
    Figure Legend Snippet: Hormonally regulated changes in the BT474 transcriptome. A Pie charts illustrating the different transcript types that are hormonally regulated in BT474 cells. In both conditions, DHT- and PROG-treated cells, 79% of the transcripts are protein-coding, 5% long non-coding RNAs (lncRNA), 2% microRNAs (miRNA), 2% small non-coding RNA (ncRNA), 2% pseudogenes, and the remaining 11% belong to other categories and gene ID duplications. *snoRNA, snRNA, scaRNA, scRNA, miscRNA. **Polymorphic_pseudogene, processed_pseudogene, transcribed_processed_pseudogene, transcribed_unitary_pseudogene, transcribed_unprocessed_ pseudogene, unitary_pseudogene, unprocessed_pseudogene. ***TEC (To be experimentally confirmed), ribosome The transcript categorization was performed on Ensembl website, using the bioinformatics tool, Biomart. B Histograms demonstrating the FC (fold change) distribution of DEGs in the transcript list, C Volcano plots illustrating the most significant differentially expressed genes. Red color shows the upregulated genes and blue color shows the downregulated genes. D Heatmap illustrating the differential expression profiles in DHT-treated and PROG-treated cells, respectively, compared to controls. Red color represents the upregulated genes, whereas the blue color represents the downregulated genes. Grey color corresponds to the genes that were not expressed, or the ratio could not be assessed in this experiment. All graphs were constructed in Perseus software using the log 2 FC values derived from the expression comparison between the hormone-treated and control cells. E , F Venn diagram representing the number and overlapping of DEGs in DHT-treated BT474 cells with known AR-regulated and cancer/BCa-related genes. The identification of the cancer/BCa related enriched genes were identified with the Gene Ontology annotation ( E ) and the KEGG analysis ( F ) of DAVID (2021 Update) online tool. G Comparison of gene set numbers identified in GSEA analysis of the RNA-seq experiment in DHT and PROG-treated BT474 cells. For the GSEA (v.3.0) analysis, only gene sets that contained 15–200 genes were used (q-value

    Techniques Used: Expressing, Construct, Software, Derivative Assay, RNA Sequencing Assay

    Schematic representation of the significant DEGs of DHT-regulated BT474 cells, using the online tool, STRING. The genes associated with either fatty acid metabolism or the WNT-pathway, cluster clearly together. Lines with different colors represent the source of data that indicate the protein–protein interaction light blue: from curated databases, purple: experimentally determined, green: gene neighborhood, red: gene fusions, dark blue: gene co-occurrence, yellow: text mining, black: co-expression, and grey: protein homology
    Figure Legend Snippet: Schematic representation of the significant DEGs of DHT-regulated BT474 cells, using the online tool, STRING. The genes associated with either fatty acid metabolism or the WNT-pathway, cluster clearly together. Lines with different colors represent the source of data that indicate the protein–protein interaction light blue: from curated databases, purple: experimentally determined, green: gene neighborhood, red: gene fusions, dark blue: gene co-occurrence, yellow: text mining, black: co-expression, and grey: protein homology

    Techniques Used: Expressing

    Pathway analysis of hormone-regulated genes identified by RNA sequencing of BT474 cells. Enrichment map of the DHT- ( A ) and PROG- ( B ) regulated genes. The analysis was performed using the Cytoscape software (GSEA 3.0, Enrichment Map plugin, q-value
    Figure Legend Snippet: Pathway analysis of hormone-regulated genes identified by RNA sequencing of BT474 cells. Enrichment map of the DHT- ( A ) and PROG- ( B ) regulated genes. The analysis was performed using the Cytoscape software (GSEA 3.0, Enrichment Map plugin, q-value

    Techniques Used: RNA Sequencing Assay, Software

    13) Product Images from "GIT1 protects against breast cancer growth through negative regulation of Notch"

    Article Title: GIT1 protects against breast cancer growth through negative regulation of Notch

    Journal: Nature Communications

    doi: 10.1038/s41467-022-28631-y

    Downregulated GIT1 expression in ER(−) breast tumours is associated with poor relapse-free survival in patients. a , b Immunostaining of GIT1 and ERα in one ER(+) and one ER(−) patient breast tumour section ( a ) and the quantitative analysis of GIT1 immunofluorescence relative to DAPI ( b ; ER(+) ( n = 45) versus ER(−) ( n = 30), P = 0.0006, t test). Scale bar, 10 μm. c Mass spectrometry data from CPTAC of the GIT1 protein levels in ER(+) and ER(−) patients (ER(+) ( n = 51) versus ER(−) ( n = 11), P = 0.024, t test). d , e , Western blots of GIT1 in various human breast cancer cells ( d ) and the quantitative analysis ( e ; n = 7 independent biological replicates, 184A1 versus BT474, P = 0.0004; MCF7, P = 0.0003; MDA-MB-134-VI, P = 0.015; MDA-MB-361, P
    Figure Legend Snippet: Downregulated GIT1 expression in ER(−) breast tumours is associated with poor relapse-free survival in patients. a , b Immunostaining of GIT1 and ERα in one ER(+) and one ER(−) patient breast tumour section ( a ) and the quantitative analysis of GIT1 immunofluorescence relative to DAPI ( b ; ER(+) ( n = 45) versus ER(−) ( n = 30), P = 0.0006, t test). Scale bar, 10 μm. c Mass spectrometry data from CPTAC of the GIT1 protein levels in ER(+) and ER(−) patients (ER(+) ( n = 51) versus ER(−) ( n = 11), P = 0.024, t test). d , e , Western blots of GIT1 in various human breast cancer cells ( d ) and the quantitative analysis ( e ; n = 7 independent biological replicates, 184A1 versus BT474, P = 0.0004; MCF7, P = 0.0003; MDA-MB-134-VI, P = 0.015; MDA-MB-361, P

    Techniques Used: Expressing, Immunostaining, Immunofluorescence, Mass Spectrometry, Western Blot, Multiple Displacement Amplification

    14) Product Images from "Effect of Inter-Domain Linker Composition on Biodistribution of ABD-Fused Affibody-Drug Conjugates Targeting HER2"

    Article Title: Effect of Inter-Domain Linker Composition on Biodistribution of ABD-Fused Affibody-Drug Conjugates Targeting HER2

    Journal: Pharmaceutics

    doi: 10.3390/pharmaceutics14030522

    Cell binding specificity, cellular processing, and interaction map of 99m Tc-labeled affibody-mcDM1 conjugates. ( A ) SKOV3, BT474 and SKBR3 cells were incubated with 99m Tc-Z HER2:2891 -(G 3 S) 3 -ABD-mcDM1, 99m Tc-Z HER2:2891 -(S 3 G) 3 -ABD-mcDM1, or 99m Tc-Z HER2:2891 -G 4 S-ABD-mcDM1 in the presence or absence of a 500-fold molar excess of the same non-labeled conjugate, blocked or non-blocked, respectively. The total cell-bound activity is shown on the Y -axis as a percentage of the total added activity. Asterisk (*) correspond to significant differences ( p
    Figure Legend Snippet: Cell binding specificity, cellular processing, and interaction map of 99m Tc-labeled affibody-mcDM1 conjugates. ( A ) SKOV3, BT474 and SKBR3 cells were incubated with 99m Tc-Z HER2:2891 -(G 3 S) 3 -ABD-mcDM1, 99m Tc-Z HER2:2891 -(S 3 G) 3 -ABD-mcDM1, or 99m Tc-Z HER2:2891 -G 4 S-ABD-mcDM1 in the presence or absence of a 500-fold molar excess of the same non-labeled conjugate, blocked or non-blocked, respectively. The total cell-bound activity is shown on the Y -axis as a percentage of the total added activity. Asterisk (*) correspond to significant differences ( p

    Techniques Used: Binding Assay, Labeling, Incubation, Activity Assay

    15) Product Images from "Ajuba Overexpression Promotes Breast Cancer Chemoresistance and Glucose Uptake through TAZ-GLUT3/Survivin Pathway"

    Article Title: Ajuba Overexpression Promotes Breast Cancer Chemoresistance and Glucose Uptake through TAZ-GLUT3/Survivin Pathway

    Journal: BioMed Research International

    doi: 10.1155/2022/3321409

    Ajuba expression and transfection efficiency in breast cancer cell lines. (a) Western blot was performed in a panel of breast cancer cell lines. The results showed that Ajuba protein was lower in MCF-10A cell line. Ajuba expression was higher in breast cancer cell lines (MCF-7, MDA-MB-468, BT474, BT549, T47D, and SK-BR-3), especially in triple-negative cell lines MDA-MB-468 and BT549. Relative protein levels were quantified using ImageJ. ∗ p
    Figure Legend Snippet: Ajuba expression and transfection efficiency in breast cancer cell lines. (a) Western blot was performed in a panel of breast cancer cell lines. The results showed that Ajuba protein was lower in MCF-10A cell line. Ajuba expression was higher in breast cancer cell lines (MCF-7, MDA-MB-468, BT474, BT549, T47D, and SK-BR-3), especially in triple-negative cell lines MDA-MB-468 and BT549. Relative protein levels were quantified using ImageJ. ∗ p

    Techniques Used: Expressing, Transfection, Western Blot, Multiple Displacement Amplification

    16) Product Images from "Neuropilin-1 is over-expressed in claudin-low breast cancer and promotes tumor progression through acquisition of stem cell characteristics and RAS/MAPK pathway activation"

    Article Title: Neuropilin-1 is over-expressed in claudin-low breast cancer and promotes tumor progression through acquisition of stem cell characteristics and RAS/MAPK pathway activation

    Journal: Breast Cancer Research : BCR

    doi: 10.1186/s13058-022-01501-7

    NRP1 is associated with the claudin-low signature in breast cancer cell lines and promotes proliferation. a Heatmap showing NRP1 mRNA expression across luminal, basal A, basal B and claudin-low human breast cancer cell lines, as well as association with core claudin-low signature genes [ 10 , 28 ]. NRP1 mRNA expression in b luminal, basal A and basal B breast cancer cell lines, C claudin-low versus non-claudin-low cell lines, and association with key claudin-low markers di claudin 3 (CLDN3) and dii vimentin across breast cancer cell lines. E Flow cytometry analysis of NRP1 expression across breast cancer cell lines representing different intrinsic subtypes and the claudin-low (CLDN low ) subtype. f Western blot analysis of NRP1 expression in BrCa cell lines including luminal A (LumA; T47D and MCF-7), luminal B (LumB; MDA-MB-361), HER2 + (BT-474, HCC1569 and SKBR3), basal (BT-20 and MDA-MB-468) and claudin-low (SUM159, MDA-MB-231 and HS578T) cells, with GAPDH loading control. g Western blot showing NRP1 expression in claudin-low cell lines (MDA-MB-231, BT549, SUM159 and HS578T) at day 3 post-transfection with NRP1 siRNA (siNRP1 [ 1 ] or siNRP1 [ 2 ]) or non-targeting control siRNA (siNT). h Cell viability of claudin-low cell lines (MDA-MB-231, BT549, SUM159 and HS578T) at 0, 1, 2, 4 and 7 days after transfection with NRP1 siRNA (siNRP1 [ 1 ] or siNRP1 [ 2 ]) relative to siNT as measured by CyQuant™ DNA quantification assay; n = 3 . Error bars represent SEM, * p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001, **** p ≤ 0.0001
    Figure Legend Snippet: NRP1 is associated with the claudin-low signature in breast cancer cell lines and promotes proliferation. a Heatmap showing NRP1 mRNA expression across luminal, basal A, basal B and claudin-low human breast cancer cell lines, as well as association with core claudin-low signature genes [ 10 , 28 ]. NRP1 mRNA expression in b luminal, basal A and basal B breast cancer cell lines, C claudin-low versus non-claudin-low cell lines, and association with key claudin-low markers di claudin 3 (CLDN3) and dii vimentin across breast cancer cell lines. E Flow cytometry analysis of NRP1 expression across breast cancer cell lines representing different intrinsic subtypes and the claudin-low (CLDN low ) subtype. f Western blot analysis of NRP1 expression in BrCa cell lines including luminal A (LumA; T47D and MCF-7), luminal B (LumB; MDA-MB-361), HER2 + (BT-474, HCC1569 and SKBR3), basal (BT-20 and MDA-MB-468) and claudin-low (SUM159, MDA-MB-231 and HS578T) cells, with GAPDH loading control. g Western blot showing NRP1 expression in claudin-low cell lines (MDA-MB-231, BT549, SUM159 and HS578T) at day 3 post-transfection with NRP1 siRNA (siNRP1 [ 1 ] or siNRP1 [ 2 ]) or non-targeting control siRNA (siNT). h Cell viability of claudin-low cell lines (MDA-MB-231, BT549, SUM159 and HS578T) at 0, 1, 2, 4 and 7 days after transfection with NRP1 siRNA (siNRP1 [ 1 ] or siNRP1 [ 2 ]) relative to siNT as measured by CyQuant™ DNA quantification assay; n = 3 . Error bars represent SEM, * p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001, **** p ≤ 0.0001

    Techniques Used: Expressing, Flow Cytometry, Western Blot, Multiple Displacement Amplification, Transfection, CyQUANT Assay

    17) Product Images from "Repurposing Tranexamic Acid as an Anticancer Agent"

    Article Title: Repurposing Tranexamic Acid as an Anticancer Agent

    Journal: Frontiers in Pharmacology

    doi: 10.3389/fphar.2021.792600

    TA decreases cancer cell viability and breast tumor growth. (A) Chemical structures of TA, N -acetylated TA (NacTA), and the DDAs tcyDTDO and dMtcyDTDO. (B) The indicated panel of cancer cell lines were treated for 72 h with different TA concentrations and cell viability was measured using the MTT assay. Results are the average of six replicates and presented as the average ±S.D. These findings are representative of three biological replicates with similar results. (C) MDA-MB-468 and BT474 cells were treated for 24 h with the indicated concentrations of TA and the incorporation of tritiated thymidine into DNA was evaluated. Results are the average of six replicates and presented as the average ±S.D. These findings are representative of three biological replicates with similar results. (D) Effects of TA on the viability of a panel of leukemia, lymphoma, and multiple myeloma cell lines. Results are the average of six replicates and presented as the average ±S.D. These findings are representative of three biological replicates with similar results. (E) Cell viability assay showing that TA more potently reduces the viability of MCF10A mammary epithelial cells overexpressing MYC compared with the vector control line. Statistically significant differences between the lines were observed at 0.25, 0.5, 1, and 2% TA. p -values were calculated using unpaired Student’s t -tests. Results are the average of six replicates and presented as the average ±S.D. These findings are representative of three biological replicates with similar results. (F) Tumor growth study in which mice bearing orthotopic 012/LVM2/LR10 mammary tumors were treated once daily with 375 mg/kg TA. The upper panel illustrates how the 012/LVM2/LR10 cancer model was derived. Tumor dimensions were measured daily and tumor volumes calculated. Averages of within-group tumor volumes are presented and error bars represent standard error of the mean. (G) Tumor volumes measured at day 12 exhibit a statistically significant TA-mediated reduction in tumor volume ( p = 0.035), with an approximately nine-fold smaller tumor volume in the TA group than the vehicle group. p -values were calculated using an unpaired Student’s t -test. Each treatment group contained five animals.
    Figure Legend Snippet: TA decreases cancer cell viability and breast tumor growth. (A) Chemical structures of TA, N -acetylated TA (NacTA), and the DDAs tcyDTDO and dMtcyDTDO. (B) The indicated panel of cancer cell lines were treated for 72 h with different TA concentrations and cell viability was measured using the MTT assay. Results are the average of six replicates and presented as the average ±S.D. These findings are representative of three biological replicates with similar results. (C) MDA-MB-468 and BT474 cells were treated for 24 h with the indicated concentrations of TA and the incorporation of tritiated thymidine into DNA was evaluated. Results are the average of six replicates and presented as the average ±S.D. These findings are representative of three biological replicates with similar results. (D) Effects of TA on the viability of a panel of leukemia, lymphoma, and multiple myeloma cell lines. Results are the average of six replicates and presented as the average ±S.D. These findings are representative of three biological replicates with similar results. (E) Cell viability assay showing that TA more potently reduces the viability of MCF10A mammary epithelial cells overexpressing MYC compared with the vector control line. Statistically significant differences between the lines were observed at 0.25, 0.5, 1, and 2% TA. p -values were calculated using unpaired Student’s t -tests. Results are the average of six replicates and presented as the average ±S.D. These findings are representative of three biological replicates with similar results. (F) Tumor growth study in which mice bearing orthotopic 012/LVM2/LR10 mammary tumors were treated once daily with 375 mg/kg TA. The upper panel illustrates how the 012/LVM2/LR10 cancer model was derived. Tumor dimensions were measured daily and tumor volumes calculated. Averages of within-group tumor volumes are presented and error bars represent standard error of the mean. (G) Tumor volumes measured at day 12 exhibit a statistically significant TA-mediated reduction in tumor volume ( p = 0.035), with an approximately nine-fold smaller tumor volume in the TA group than the vehicle group. p -values were calculated using an unpaired Student’s t -test. Each treatment group contained five animals.

    Techniques Used: MTT Assay, Multiple Displacement Amplification, Viability Assay, Plasmid Preparation, Mouse Assay, Derivative Assay

    18) Product Images from "BHMPS Inhibits Breast Cancer Migration and Invasion by Disrupting Rab27a-Mediated EGFR and Fibronectin Secretion"

    Article Title: BHMPS Inhibits Breast Cancer Migration and Invasion by Disrupting Rab27a-Mediated EGFR and Fibronectin Secretion

    Journal: Cancers

    doi: 10.3390/cancers14020373

    Rab27a overexpression promotes breast cancer cell migration. ( A ) BT474 and MCF7 cells were transfected with control or Rab27a expression vector. After 24 h, the cells were treated with 10 µM of BHMPS, and the percentage of migrated cells was determined. Statistically significant differences are shown in comparison with the Rab27a overexpression group. Representative images (40× magnification, Scale bars 250 µm) and means ± SD are from three independent experiments. ** p
    Figure Legend Snippet: Rab27a overexpression promotes breast cancer cell migration. ( A ) BT474 and MCF7 cells were transfected with control or Rab27a expression vector. After 24 h, the cells were treated with 10 µM of BHMPS, and the percentage of migrated cells was determined. Statistically significant differences are shown in comparison with the Rab27a overexpression group. Representative images (40× magnification, Scale bars 250 µm) and means ± SD are from three independent experiments. ** p

    Techniques Used: Over Expression, Migration, Transfection, Expressing, Plasmid Preparation

    19) Product Images from "Building a Foundation for Precision Onco-Nutrition: Docosahexaenoic Acid and Breast Cancer"

    Article Title: Building a Foundation for Precision Onco-Nutrition: Docosahexaenoic Acid and Breast Cancer

    Journal: Cancers

    doi: 10.3390/cancers14010157

    Dose- and time-dependent effects of n6- or n3-fatty acids on the growth of five subtype human breast cancer cells (BT474, MCF7, SKBR3, MDAMB231, and MDAMB468) graphed using the area under the curve (AUC) as a percent of the untreated control. ( a ) Linoleic acid (LA, n6); ( b ) Eicosapentaenoic acid (EPA, n3); ( c ) Docosahexaenoic acid (DHA, n3).
    Figure Legend Snippet: Dose- and time-dependent effects of n6- or n3-fatty acids on the growth of five subtype human breast cancer cells (BT474, MCF7, SKBR3, MDAMB231, and MDAMB468) graphed using the area under the curve (AUC) as a percent of the untreated control. ( a ) Linoleic acid (LA, n6); ( b ) Eicosapentaenoic acid (EPA, n3); ( c ) Docosahexaenoic acid (DHA, n3).

    Techniques Used:

    Effect of 4-oxo-DHA on Cell Transcription Factors. Levels of cell proliferation or apoptosis and associated target proteins in three human breast cancer cell lines (BT474, SKBR3, and MDAMB468); ( a ) orthogonal projections to latent structures-discriminant analysis (OPLS-DA) shows a 3-class supervised model and partition the sources of variation; ( b ) to visualize the misclassification rate, the dendrogram depicts hierarchical clustering patterns among three different cell lines.
    Figure Legend Snippet: Effect of 4-oxo-DHA on Cell Transcription Factors. Levels of cell proliferation or apoptosis and associated target proteins in three human breast cancer cell lines (BT474, SKBR3, and MDAMB468); ( a ) orthogonal projections to latent structures-discriminant analysis (OPLS-DA) shows a 3-class supervised model and partition the sources of variation; ( b ) to visualize the misclassification rate, the dendrogram depicts hierarchical clustering patterns among three different cell lines.

    Techniques Used:

    Metabolism of docosahexaenoic acid (DHA) and the dose- and time-dependent effects of 4-oxo-DHA on the growth of five subtype human breast cancer cells (BT474, MCF7, SKBR3, MDAMB231, and MDAMB468) graphed using an area under the curve (AUC) or the half-maximal inhibitory concentration (IC 50 ) as a percent of untreated control. ( a ) Metabolism of DHA; ( b ) an area under the curve analysis; ( c ) the half-maximal inhibitory concentration for 3 days of treatment.
    Figure Legend Snippet: Metabolism of docosahexaenoic acid (DHA) and the dose- and time-dependent effects of 4-oxo-DHA on the growth of five subtype human breast cancer cells (BT474, MCF7, SKBR3, MDAMB231, and MDAMB468) graphed using an area under the curve (AUC) or the half-maximal inhibitory concentration (IC 50 ) as a percent of untreated control. ( a ) Metabolism of DHA; ( b ) an area under the curve analysis; ( c ) the half-maximal inhibitory concentration for 3 days of treatment.

    Techniques Used: Concentration Assay

    Canonical pathways affected by 4-oxo-DHA across three molecular subtypes of breast cancer cell lines. Canonical pathways affected by 4-oxo-DHA as predicted by the nonphosphorylated proteome data sets listed in Table 1 , Table 2 , Table 3 and Table 4 . Canonical pathways shown for cell line ( a ) BT474, ( b ) MDAMB468, and ( c ) SKBR3.The −log(B-H p -values) is the multicomparison adjusted probability that the association between the protein expression set and the canonical pathway is due to chance. The direction of the differences in expression between the treatment and the control for each protein component was compared to that tabulated in the IPA knowledge basis ( > 80,000 database entries) that support the canonical pathways that have been annotated and a z-score was computed. A z-score ≤ −2 indicates the pathway is inhibited and ≥2 that the pathway is activated. If the z-score is between −2 and 2, no prediction of inhibition or activation is deduced. Shades of red indicate that the pathway was activated by treatment with 4-oxo-DHA; greater color intensity indicates stronger evidence of activation. Shades of blue indicate that the pathway was inhibited by treatment with 4-oxo-DHA; greater color intensity indicates stronger evidence of inhibition. The absence of coloration indicates that the evidence was not strong enough to permit a prediction of pathway status.
    Figure Legend Snippet: Canonical pathways affected by 4-oxo-DHA across three molecular subtypes of breast cancer cell lines. Canonical pathways affected by 4-oxo-DHA as predicted by the nonphosphorylated proteome data sets listed in Table 1 , Table 2 , Table 3 and Table 4 . Canonical pathways shown for cell line ( a ) BT474, ( b ) MDAMB468, and ( c ) SKBR3.The −log(B-H p -values) is the multicomparison adjusted probability that the association between the protein expression set and the canonical pathway is due to chance. The direction of the differences in expression between the treatment and the control for each protein component was compared to that tabulated in the IPA knowledge basis ( > 80,000 database entries) that support the canonical pathways that have been annotated and a z-score was computed. A z-score ≤ −2 indicates the pathway is inhibited and ≥2 that the pathway is activated. If the z-score is between −2 and 2, no prediction of inhibition or activation is deduced. Shades of red indicate that the pathway was activated by treatment with 4-oxo-DHA; greater color intensity indicates stronger evidence of activation. Shades of blue indicate that the pathway was inhibited by treatment with 4-oxo-DHA; greater color intensity indicates stronger evidence of inhibition. The absence of coloration indicates that the evidence was not strong enough to permit a prediction of pathway status.

    Techniques Used: Expressing, Indirect Immunoperoxidase Assay, Inhibition, Activation Assay

    Effect of 4-oxo-DHA on the mTOR signaling pathway was diagrammed in IPA and corresponding expression data from the BT474 cell line was overlaid on the pathway. Overlaid expression is shown as the ratio of phosphorylated to total protein. The chart on the right shows the expression of major regulatory nodes of the mTOR signaling pathway while illustrating a pattern of differential regulation across the cell lines BT474, SRBR3, and MDAMB464, respectively. Red indicates protein activation and blue indicates inhibition.
    Figure Legend Snippet: Effect of 4-oxo-DHA on the mTOR signaling pathway was diagrammed in IPA and corresponding expression data from the BT474 cell line was overlaid on the pathway. Overlaid expression is shown as the ratio of phosphorylated to total protein. The chart on the right shows the expression of major regulatory nodes of the mTOR signaling pathway while illustrating a pattern of differential regulation across the cell lines BT474, SRBR3, and MDAMB464, respectively. Red indicates protein activation and blue indicates inhibition.

    Techniques Used: Indirect Immunoperoxidase Assay, Expressing, Activation Assay, Inhibition

    Effect of 4-oxo-DHA on Growth Factor Signaling. Levels of cell proliferation or apoptosis and associated target proteins in three human breast cancer cell lines (BT474, SKBR3, and MDAMB468); ( a ) Orthogonal projections to latent structures-discriminant analysis (OPLS-DA) shows a 3-class supervised model and partition the sources of variation; ( b ) To visualize the misclassification rate, the dendrogram depicts hierarchical clustering patterns among three different cell lines.
    Figure Legend Snippet: Effect of 4-oxo-DHA on Growth Factor Signaling. Levels of cell proliferation or apoptosis and associated target proteins in three human breast cancer cell lines (BT474, SKBR3, and MDAMB468); ( a ) Orthogonal projections to latent structures-discriminant analysis (OPLS-DA) shows a 3-class supervised model and partition the sources of variation; ( b ) To visualize the misclassification rate, the dendrogram depicts hierarchical clustering patterns among three different cell lines.

    Techniques Used:

    Canonical pathways affected by 4-oxo-DHA as predicted by the phosphoproteome data sets listed in Table 1 , Table 2 , Table 3 and Table 4 . Canonical pathways shown for cell line ( a ) BT474, ( b ) MDAMB468, and ( c ) SKBR3. The −log(B-H p -values) is the multi = comparison-adjusted probability that the association between the protein expression set and the canonical pathway is due to chance. The direction of the differences in expression between the treatment and the control for each protein component was compared to that tabulated in the IPA knowledge basis ( > 80,000 database entries) that support the canonical pathways that have been annotated and a z-score was computed. A z-score ≤ −2 indicates the pathway is inhibited and ≥2 that the pathway is activated. If the z-score is between −2 and 2, no prediction of inhibition or activation is deduced. Shades of red indicate that the pathway was activated by treatment with 4-oxo-DHA; greater color intensity indicates stronger evidence of activation. Shades of blue indicate that the pathway was inhibited by treatment with 4-oxo-DHA; greater color intensity indicates stronger evidence of inhibition. The absence of coloration indicates that the evidence was not strong enough to permit a prediction of pathway status.
    Figure Legend Snippet: Canonical pathways affected by 4-oxo-DHA as predicted by the phosphoproteome data sets listed in Table 1 , Table 2 , Table 3 and Table 4 . Canonical pathways shown for cell line ( a ) BT474, ( b ) MDAMB468, and ( c ) SKBR3. The −log(B-H p -values) is the multi = comparison-adjusted probability that the association between the protein expression set and the canonical pathway is due to chance. The direction of the differences in expression between the treatment and the control for each protein component was compared to that tabulated in the IPA knowledge basis ( > 80,000 database entries) that support the canonical pathways that have been annotated and a z-score was computed. A z-score ≤ −2 indicates the pathway is inhibited and ≥2 that the pathway is activated. If the z-score is between −2 and 2, no prediction of inhibition or activation is deduced. Shades of red indicate that the pathway was activated by treatment with 4-oxo-DHA; greater color intensity indicates stronger evidence of activation. Shades of blue indicate that the pathway was inhibited by treatment with 4-oxo-DHA; greater color intensity indicates stronger evidence of inhibition. The absence of coloration indicates that the evidence was not strong enough to permit a prediction of pathway status.

    Techniques Used: Expressing, Indirect Immunoperoxidase Assay, Inhibition, Activation Assay

    Effect of 4-oxo-DHA on Cell Proliferation and Apoptotic Cell Death. Levels of cell proliferation or apoptosis and associated target proteins in three human breast cancer cell lines (BT474, SKBR3, and MDAMB468); ( a ) Orthogonal projections to latent structures-discriminant analysis (OPLS-DA) shows a 3-class supervised model and partition the sources of variation; ( b ) To visualize the misclassification rate, the dendrogram depicts hierarchical clustering patterns among three different cell lines.
    Figure Legend Snippet: Effect of 4-oxo-DHA on Cell Proliferation and Apoptotic Cell Death. Levels of cell proliferation or apoptosis and associated target proteins in three human breast cancer cell lines (BT474, SKBR3, and MDAMB468); ( a ) Orthogonal projections to latent structures-discriminant analysis (OPLS-DA) shows a 3-class supervised model and partition the sources of variation; ( b ) To visualize the misclassification rate, the dendrogram depicts hierarchical clustering patterns among three different cell lines.

    Techniques Used:

    20) Product Images from "MAL2 mediates the formation of stable HER2 signaling complexes within lipid raft-rich membrane protrusions in breast cancer cells"

    Article Title: MAL2 mediates the formation of stable HER2 signaling complexes within lipid raft-rich membrane protrusions in breast cancer cells

    Journal: Cell reports

    doi: 10.1016/j.celrep.2021.110160

    Enrichment of PIP2 and pAKT together with HER2 in membrane protrusions requires MAL2 and lipid rafts (A) Co-immunofluorescence staining for Ezrin (top) or HER2 (bottom) with phalloidin in PH-PLCδ-GFP-expressing control SKBR3 cells. Scale bars represent 10 μm. (B) Immunofluorescence staining for Ezrin (top) or HER2 (bottom) with phalloidin in PH-PLCδ-GFP-expressing MAL2KD_SKBR3 cells. (C) Immunofluorescence staining for Ezrin (top) or HER2 (bottom) with phalloidin in PH-PLCδ-GFP-expressing SKBR3 cells treated with MβCD. (D) Immunofluorescence staining for Ezrin with phalloidin in PH-PLCδ-GFP-expressing SKBR3 cells treated with wortmannin (0, 5, and 10 μM). (E) Immunofluorescence staining for HER2 with phalloidin in PH-PLCδ-GFP-expressing SKBR3 cells treated with wortmannin (0, 5, and 10 μM). (F) PLA results for HER2 and Ezrin in control and wortmannin-treated SKBR3 cells. Scale bars represent 10 μm. (G) Western blot analysis of AKT and phospho-AKT in control and MAL2 knockdown SKBR3 cells (left) and in control and MβCD-treated SKBR3 cells (right). (H–J) SIM imaging showing HER2, pAKT, and phalloidin immunofluorescence in control (H), MAL2KD (I)-treated, and MβCD-treated (J) SKBR3 cells. Scale bars represent 10 μm. (K) Immunofluorescence staining for FOXO1 in control, MAL2KD-treated, and MβCD-treated SKBR3 cells. (L) XTT cell viability assay in control, MAL2KD-treated, and MβCD-treated SKBR3 and BT474 cells. Bar graphs represent the mean ± SEM. ****p
    Figure Legend Snippet: Enrichment of PIP2 and pAKT together with HER2 in membrane protrusions requires MAL2 and lipid rafts (A) Co-immunofluorescence staining for Ezrin (top) or HER2 (bottom) with phalloidin in PH-PLCδ-GFP-expressing control SKBR3 cells. Scale bars represent 10 μm. (B) Immunofluorescence staining for Ezrin (top) or HER2 (bottom) with phalloidin in PH-PLCδ-GFP-expressing MAL2KD_SKBR3 cells. (C) Immunofluorescence staining for Ezrin (top) or HER2 (bottom) with phalloidin in PH-PLCδ-GFP-expressing SKBR3 cells treated with MβCD. (D) Immunofluorescence staining for Ezrin with phalloidin in PH-PLCδ-GFP-expressing SKBR3 cells treated with wortmannin (0, 5, and 10 μM). (E) Immunofluorescence staining for HER2 with phalloidin in PH-PLCδ-GFP-expressing SKBR3 cells treated with wortmannin (0, 5, and 10 μM). (F) PLA results for HER2 and Ezrin in control and wortmannin-treated SKBR3 cells. Scale bars represent 10 μm. (G) Western blot analysis of AKT and phospho-AKT in control and MAL2 knockdown SKBR3 cells (left) and in control and MβCD-treated SKBR3 cells (right). (H–J) SIM imaging showing HER2, pAKT, and phalloidin immunofluorescence in control (H), MAL2KD (I)-treated, and MβCD-treated (J) SKBR3 cells. Scale bars represent 10 μm. (K) Immunofluorescence staining for FOXO1 in control, MAL2KD-treated, and MβCD-treated SKBR3 cells. (L) XTT cell viability assay in control, MAL2KD-treated, and MβCD-treated SKBR3 and BT474 cells. Bar graphs represent the mean ± SEM. ****p

    Techniques Used: Immunofluorescence, Staining, Expressing, Proximity Ligation Assay, Western Blot, Imaging, Viability Assay

    MAL2 expression is required for formation of lipid raft-enriched membrane protrusions (A) Immunofluorescence staining for HER2 and MAL2 (top row) and MAL2 and cholera toxin B (lipid rafts) (bottom row) in SKBR3 cells. (B) Immunofluorescence staining for FLAG-tagged MAL2 or endogenous MAL2 with HER2 and actin in control and MβCD (5 mM)-treated SKBR3 cells. Scale bars represent 10 μm. (C) Proximity ligation assay (PLA) experiment for HER2 and MAL2 in control and MβCD (5 mM)-treated SKBR3 cells also stained for actin (phalloidin). Scale bars represent 10 μm. (D) MAL2 mRNA expression in control and MAL2 knockdown SKBR3 and BT474 cells as assessed by quantitative RT-PCR (n = 3). (E) Western blot analysis of HER2, phospho-HER2, EGFR, and phospho-EGFR in control and MAL2 knockdown SKBR3 and BT474 cells. (F) BrdU incorporation in MAL2KD cells relative to control SKBR3 and BT474 cells. (G) PLA experiment for HER2 and MAL2 in control and MAL2LD SKBR3 cells. Scale bars represent 10 μm. (H) Transmission electron microscopy images in control and MAL2 knockdown SKBR3 cells. (I) Immunofluorescence staining HER2 and cholera toxin B in control (top), MAL2KD-treated (middle), and MβCD-treated (5 mM) SKBR3 cells. Scale bars represent 10 μm. (J) Immunofluorescence staining for FLAG-tagged FLOT1, HER2, and phalloidin in control, MAL2KD-treated, and MβCD (5 mM)-treated SKBR3 cells. Scale bars represent 10 μm. (K) Immunofluorescence staining for HER2 and EGFR in control and MAL2KD SKBR3 cells. Scale bars represent 10 μm. (L) PLA experiment for HER2 and EGFR in control and MAL2KD SKBR3 cells also stained for actin (phalloidin). Scale bars represent 10μm. (M) Quantitation of PLA experiment by measuring fluorescent intensity of amplified PLA reactions and phalloidin. Bar graphs represent the mean ± SEM. **p
    Figure Legend Snippet: MAL2 expression is required for formation of lipid raft-enriched membrane protrusions (A) Immunofluorescence staining for HER2 and MAL2 (top row) and MAL2 and cholera toxin B (lipid rafts) (bottom row) in SKBR3 cells. (B) Immunofluorescence staining for FLAG-tagged MAL2 or endogenous MAL2 with HER2 and actin in control and MβCD (5 mM)-treated SKBR3 cells. Scale bars represent 10 μm. (C) Proximity ligation assay (PLA) experiment for HER2 and MAL2 in control and MβCD (5 mM)-treated SKBR3 cells also stained for actin (phalloidin). Scale bars represent 10 μm. (D) MAL2 mRNA expression in control and MAL2 knockdown SKBR3 and BT474 cells as assessed by quantitative RT-PCR (n = 3). (E) Western blot analysis of HER2, phospho-HER2, EGFR, and phospho-EGFR in control and MAL2 knockdown SKBR3 and BT474 cells. (F) BrdU incorporation in MAL2KD cells relative to control SKBR3 and BT474 cells. (G) PLA experiment for HER2 and MAL2 in control and MAL2LD SKBR3 cells. Scale bars represent 10 μm. (H) Transmission electron microscopy images in control and MAL2 knockdown SKBR3 cells. (I) Immunofluorescence staining HER2 and cholera toxin B in control (top), MAL2KD-treated (middle), and MβCD-treated (5 mM) SKBR3 cells. Scale bars represent 10 μm. (J) Immunofluorescence staining for FLAG-tagged FLOT1, HER2, and phalloidin in control, MAL2KD-treated, and MβCD (5 mM)-treated SKBR3 cells. Scale bars represent 10 μm. (K) Immunofluorescence staining for HER2 and EGFR in control and MAL2KD SKBR3 cells. Scale bars represent 10 μm. (L) PLA experiment for HER2 and EGFR in control and MAL2KD SKBR3 cells also stained for actin (phalloidin). Scale bars represent 10μm. (M) Quantitation of PLA experiment by measuring fluorescent intensity of amplified PLA reactions and phalloidin. Bar graphs represent the mean ± SEM. **p

    Techniques Used: Expressing, Immunofluorescence, Staining, Proximity Ligation Assay, Quantitative RT-PCR, Western Blot, BrdU Incorporation Assay, Transmission Assay, Electron Microscopy, Quantitation Assay, Amplification

    Enhanced interactions between HER2 and MAL2, Ezrin, NHERF1, and PMCA2 in trastuzumab-resistant SKBR3 cells (A) Immunofluorescence staining for cholera toxin B (lipid rafts) in control and trastuzumab-resistant S KBR3 cells. Scale bars represent 10 μm. (B) Immunofluorescence staining for HER2 and MAL2 in control and trastuzumab-resistant SKBR3 cells. Scale bars represent 10 μm. (C) PLA for HER2 and MAL2 in control and trastuzumab-resistant SKBR3 cells also stained for phalloidin. Scale bars represent 10 μm. (D–F) Quantitative results from immunoprecipitation coupled with data-independent acquisition mass spectrometry (DIA-MS) in control and trastuzumab-resistant SKBR3 cells. (D) The DIA-MS Intensity (log 2 ) of HER2 and MAL2 proteins from control and trastuzumab-resistant SKBR3 cells. (E) The DIA-MS Intensity (log 2 ) for all the peptide precursor signals of HER2 in control and resistant cells. (F) The DIA-MS peak groups visualized for quantifying HER2 (VLGSGAFGTVYK) and MAL2 (VTLPAGPDILR). Peaks above and below the middle line denote the MS2 and MS1 ion traces in DIA-MS. (G) MAL2, Ezrin, and NHERF1 mRNA expression in control and trastuzumab-resistant SKBR3 cells as assessed by quantitative RT-PCR (n = 3). (H) PLA for HER2 with Ezrin (left), NHERF1 (middle), and PMCA2 (right) in control and trastuzumab-resistant SKBR3 cells also stained for phalloidin. Boxed portions are amplified at right with co-registration of PLA signal and immunofluorescence for actin (phalloidin). (I) Quantitation of PLA experiment for HER2 in combination with MAL2, Ezrin, NHERF1, or PMCA2 represented as the fluorescent intensity of amplified PLA signals associated with membrane protrusions. (J) Coimmunoprecipitation for HER2 and HSP90 in control and trastuzumab-resistant SKBR3 cells. (K) PLA for HER2 and HSP90 in control and trastuzumab-resistant SKBR3 cells also stained for phalloidin. Scale bars represent 10 μm. (L) XTT cell viability assay in control, MAL2KD-treated, and MβCD-treated trastuzumab-resistant SKBR3 and BT474 cells. (M) Immunofluorescence staining for FOXO1 in control, MAL2KD-treated, and MβCD-treated trastuzumab-resistant SKBR3 cells. (N) Immunofluorescence staining for HER2 and pAKT in control, MAL2KD-treated, and MβCD-treated trastuzumab-resistant SKBR3 cells. Scale bars represent 10 μm. (O) Diagram representing the structure of MAL2- and lipid raft-enriched membrane protrusions containing multi-protein HER2 signaling complexes. These results are representative of three independent experiments.
    Figure Legend Snippet: Enhanced interactions between HER2 and MAL2, Ezrin, NHERF1, and PMCA2 in trastuzumab-resistant SKBR3 cells (A) Immunofluorescence staining for cholera toxin B (lipid rafts) in control and trastuzumab-resistant S KBR3 cells. Scale bars represent 10 μm. (B) Immunofluorescence staining for HER2 and MAL2 in control and trastuzumab-resistant SKBR3 cells. Scale bars represent 10 μm. (C) PLA for HER2 and MAL2 in control and trastuzumab-resistant SKBR3 cells also stained for phalloidin. Scale bars represent 10 μm. (D–F) Quantitative results from immunoprecipitation coupled with data-independent acquisition mass spectrometry (DIA-MS) in control and trastuzumab-resistant SKBR3 cells. (D) The DIA-MS Intensity (log 2 ) of HER2 and MAL2 proteins from control and trastuzumab-resistant SKBR3 cells. (E) The DIA-MS Intensity (log 2 ) for all the peptide precursor signals of HER2 in control and resistant cells. (F) The DIA-MS peak groups visualized for quantifying HER2 (VLGSGAFGTVYK) and MAL2 (VTLPAGPDILR). Peaks above and below the middle line denote the MS2 and MS1 ion traces in DIA-MS. (G) MAL2, Ezrin, and NHERF1 mRNA expression in control and trastuzumab-resistant SKBR3 cells as assessed by quantitative RT-PCR (n = 3). (H) PLA for HER2 with Ezrin (left), NHERF1 (middle), and PMCA2 (right) in control and trastuzumab-resistant SKBR3 cells also stained for phalloidin. Boxed portions are amplified at right with co-registration of PLA signal and immunofluorescence for actin (phalloidin). (I) Quantitation of PLA experiment for HER2 in combination with MAL2, Ezrin, NHERF1, or PMCA2 represented as the fluorescent intensity of amplified PLA signals associated with membrane protrusions. (J) Coimmunoprecipitation for HER2 and HSP90 in control and trastuzumab-resistant SKBR3 cells. (K) PLA for HER2 and HSP90 in control and trastuzumab-resistant SKBR3 cells also stained for phalloidin. Scale bars represent 10 μm. (L) XTT cell viability assay in control, MAL2KD-treated, and MβCD-treated trastuzumab-resistant SKBR3 and BT474 cells. (M) Immunofluorescence staining for FOXO1 in control, MAL2KD-treated, and MβCD-treated trastuzumab-resistant SKBR3 cells. (N) Immunofluorescence staining for HER2 and pAKT in control, MAL2KD-treated, and MβCD-treated trastuzumab-resistant SKBR3 cells. Scale bars represent 10 μm. (O) Diagram representing the structure of MAL2- and lipid raft-enriched membrane protrusions containing multi-protein HER2 signaling complexes. These results are representative of three independent experiments.

    Techniques Used: Immunofluorescence, Staining, Proximity Ligation Assay, Immunoprecipitation, Mass Spectrometry, Expressing, Quantitative RT-PCR, Amplification, Quantitation Assay, Viability Assay

    21) Product Images from "Transforming Tea Catechins into Potent Anticancer Compound: Analysis of Three Boronated-PEG Delivery System"

    Article Title: Transforming Tea Catechins into Potent Anticancer Compound: Analysis of Three Boronated-PEG Delivery System

    Journal: Micromachines

    doi: 10.3390/mi13010045

    Cellular uptake of control ( A ), rhodamine encapsulated NP(EGCG) ( B ); NP (PIC) ( C ); and NP(EPI) ( D ) in BT474 cells in vitro.
    Figure Legend Snippet: Cellular uptake of control ( A ), rhodamine encapsulated NP(EGCG) ( B ); NP (PIC) ( C ); and NP(EPI) ( D ) in BT474 cells in vitro.

    Techniques Used: In Vitro

    22) Product Images from "The Synergistic Effects of SHR6390 Combined With Pyrotinib on HER2+/HR+ Breast Cancer"

    Article Title: The Synergistic Effects of SHR6390 Combined With Pyrotinib on HER2+/HR+ Breast Cancer

    Journal: Frontiers in Cell and Developmental Biology

    doi: 10.3389/fcell.2021.785796

    Molecular mechanism studies in HER2+/HR+ breast cancer cell lines after treatment and in vivo anticancer effect of PYR and SHR in cancer xenograft models. EFM-192A cells were treated with PBS, PBS (CON), PYR (0.16, 0.4 μg/ml), SHR (0.6, 1.5 μg/ml) or PYR (0.4 μg/ml), and SHR (1.5 μg/ml) for 24 h, respectively. BT474 cells were treated with PBS, PYR (0.96, 2.4 μg/ml), SHR (1.2, 3 μg/ml) or PYR (2.4 μg/ml), and SHR (3 μg/ml) for 24 h, respectively. Nuclear and cytosolic protein extracts were subjected to Western blot analysis. (5A,5B) The results of Western blot for FOXM1, pFOXM1, NFκB, pNFκB, GSK3β, pGSK3β, RB, and pRB in the nuclear fractions and cytosolic extracts, respectively. GAPDH served as the loading control. (5C,5D) Quantitative analysis of the Western blotting results. Data represent the mean ±S.D. of three independent experiments. Randomly grouped nude mice were treated with PBS (Vehicle), PYR (10 mg/kg/day), SHR (75 mg/kg/day), or a combination treatment (PYR+SHR) for 25 days. (5E) Photos of the excised tumors. (5F,5G) Tumor growth ratio curve and body weight changes every 3 days after the onset of treatment. (5H) Weight obtained on day 25 after treatment, * p
    Figure Legend Snippet: Molecular mechanism studies in HER2+/HR+ breast cancer cell lines after treatment and in vivo anticancer effect of PYR and SHR in cancer xenograft models. EFM-192A cells were treated with PBS, PBS (CON), PYR (0.16, 0.4 μg/ml), SHR (0.6, 1.5 μg/ml) or PYR (0.4 μg/ml), and SHR (1.5 μg/ml) for 24 h, respectively. BT474 cells were treated with PBS, PYR (0.96, 2.4 μg/ml), SHR (1.2, 3 μg/ml) or PYR (2.4 μg/ml), and SHR (3 μg/ml) for 24 h, respectively. Nuclear and cytosolic protein extracts were subjected to Western blot analysis. (5A,5B) The results of Western blot for FOXM1, pFOXM1, NFκB, pNFκB, GSK3β, pGSK3β, RB, and pRB in the nuclear fractions and cytosolic extracts, respectively. GAPDH served as the loading control. (5C,5D) Quantitative analysis of the Western blotting results. Data represent the mean ±S.D. of three independent experiments. Randomly grouped nude mice were treated with PBS (Vehicle), PYR (10 mg/kg/day), SHR (75 mg/kg/day), or a combination treatment (PYR+SHR) for 25 days. (5E) Photos of the excised tumors. (5F,5G) Tumor growth ratio curve and body weight changes every 3 days after the onset of treatment. (5H) Weight obtained on day 25 after treatment, * p

    Techniques Used: In Vivo, Western Blot, Mouse Assay

    Effects of PYR and SHR on HER2+/HR+ breast cancer cell migration and invasion. (3A) Wound healing assay assessed the effect of PYR (0.16, 0.4 μg/ml), SHR (0.6, 1.5 μg/ml), or PYR (0.4 μg/ml) and SHR (1.5 μg/ml) on EFM-192A cells’ migration ability, and (3B) histogram represents the statistical analysis. (3C) Wound healing assay assessed the effect of PYR (0.96, 2.4 μg/ml), SHR (1.2, 3 μg/ml) or PYR (2.4 μg/ml), and SHR (3 μg/ml) on BT474 cells’ migration ability, and (3D) histogram represents the statistical analysis. Cell invasion was analyzed with a Matrigel-coated Boyden chamber. HER2+/HR+ breast cancer cell lines were treated with PBS(CON) and different concentrations of PYR and SHR for 24 h. (3E) Transwell invasion assays assessed the effect of PYR (0.16, 0.4 μg/ml), SHR (0.6, 1.5 μg/ml) or PYR (0.4 μg/ml), and SHR (1.5 μg/ml) on EFM-192A cell invasion ability, and (3F) histogram represents the statistical analysis. (3G) Transwell invasion assays assessed the effect of PYR (0.96, 2.4 μg/ml), SHR (1.2, 3 μg/ml) or PYR (2.4 μg/ml), and SHR (3 μg/ml) on BT474 cells’ invasion ability, and (3H) histogram represents the statistical analysis. Original magnification was ×100. Data represent the mean ± S.D. of three independent experiments. * p
    Figure Legend Snippet: Effects of PYR and SHR on HER2+/HR+ breast cancer cell migration and invasion. (3A) Wound healing assay assessed the effect of PYR (0.16, 0.4 μg/ml), SHR (0.6, 1.5 μg/ml), or PYR (0.4 μg/ml) and SHR (1.5 μg/ml) on EFM-192A cells’ migration ability, and (3B) histogram represents the statistical analysis. (3C) Wound healing assay assessed the effect of PYR (0.96, 2.4 μg/ml), SHR (1.2, 3 μg/ml) or PYR (2.4 μg/ml), and SHR (3 μg/ml) on BT474 cells’ migration ability, and (3D) histogram represents the statistical analysis. Cell invasion was analyzed with a Matrigel-coated Boyden chamber. HER2+/HR+ breast cancer cell lines were treated with PBS(CON) and different concentrations of PYR and SHR for 24 h. (3E) Transwell invasion assays assessed the effect of PYR (0.16, 0.4 μg/ml), SHR (0.6, 1.5 μg/ml) or PYR (0.4 μg/ml), and SHR (1.5 μg/ml) on EFM-192A cell invasion ability, and (3F) histogram represents the statistical analysis. (3G) Transwell invasion assays assessed the effect of PYR (0.96, 2.4 μg/ml), SHR (1.2, 3 μg/ml) or PYR (2.4 μg/ml), and SHR (3 μg/ml) on BT474 cells’ invasion ability, and (3H) histogram represents the statistical analysis. Original magnification was ×100. Data represent the mean ± S.D. of three independent experiments. * p

    Techniques Used: Migration, Wound Healing Assay

    Effects of PYR and SHR on the cell apoptosis and cell cycle. (4A) Cell cycle analysis through PI staining and following flow cytometry for EFM-192A cells after incubated with PBS (CON), PYR (0.16, 0.4 μg/ml), SHR (0.6, 1.5 μg/ml) or PYR (0.4 μg/ml), and SHR (1.5 μg/ml) for 24 h. ModFit was used to perform cell cycle analysis. (4B) Cell cycle analysis through PI staining and following flow cytometry for BT474 cells after being incubated with PBS (CON), PYR (0.96, 2.4 μg/ml), SHR (1.2, 3 μg/ml) or PYR (2.4 μg/ml), and SHR (3 μg/ml) for 24 h. (4E,4F) The histograms were the representative results. Compared with control, PYR and SHR caused significant G1/S phase arrest. (4C) Cell apoptosis was detected through Annexin V-FITC/PI double staining and following flow cytometry for EFM-192A cells after being incubated with PBS (CON), PYR (0.16, 0.4 μg/ml), SHR (0.6, 1.5 μg/ml) or PYR (0.4 μg/ml), and SHR (1.5 μg/ml) for 24 h. (4D) Cell apoptosis for BT474 cells after being incubated with PBS (CON), PYR (0.96, 2.4 μg/ml), SHR (1.2, 3 μg/ml) or PYR (2.4 μg/ml), and SHR (3 μg/ml). (4G,4H) The histograms were the representative results. Compared to the control, PRY and SHR promote apoptosis and induces cellular stress in HER2+/HR+ breast cancer cell lines. Data represent the mean ± S.D. of three independent experiments.
    Figure Legend Snippet: Effects of PYR and SHR on the cell apoptosis and cell cycle. (4A) Cell cycle analysis through PI staining and following flow cytometry for EFM-192A cells after incubated with PBS (CON), PYR (0.16, 0.4 μg/ml), SHR (0.6, 1.5 μg/ml) or PYR (0.4 μg/ml), and SHR (1.5 μg/ml) for 24 h. ModFit was used to perform cell cycle analysis. (4B) Cell cycle analysis through PI staining and following flow cytometry for BT474 cells after being incubated with PBS (CON), PYR (0.96, 2.4 μg/ml), SHR (1.2, 3 μg/ml) or PYR (2.4 μg/ml), and SHR (3 μg/ml) for 24 h. (4E,4F) The histograms were the representative results. Compared with control, PYR and SHR caused significant G1/S phase arrest. (4C) Cell apoptosis was detected through Annexin V-FITC/PI double staining and following flow cytometry for EFM-192A cells after being incubated with PBS (CON), PYR (0.16, 0.4 μg/ml), SHR (0.6, 1.5 μg/ml) or PYR (0.4 μg/ml), and SHR (1.5 μg/ml) for 24 h. (4D) Cell apoptosis for BT474 cells after being incubated with PBS (CON), PYR (0.96, 2.4 μg/ml), SHR (1.2, 3 μg/ml) or PYR (2.4 μg/ml), and SHR (3 μg/ml). (4G,4H) The histograms were the representative results. Compared to the control, PRY and SHR promote apoptosis and induces cellular stress in HER2+/HR+ breast cancer cell lines. Data represent the mean ± S.D. of three independent experiments.

    Techniques Used: Cell Cycle Assay, Staining, Flow Cytometry, Incubation, Double Staining

    Effects of PYR and SHR on the viability of HER2+/HR+ breast cancer cell lines. (1A) Proliferation activity of the EFM-192A cell line was determined by the MTT assay after incubation for 48 h with different concentrations of PYR or SHR. (1B – 1D) The EFM-192A cell line was treated with PYR or SHR alone or in combination or in sequences (PYR first for 6 h followed by SHR or SHR first for 6 h followed by PYR). (1E) Proliferation activity of the EFM-192A cell line was determined after incubation for 48 h with different concentrations of SHR combination with 0.1875, 0.375, 0.75, 1.5, and 3 μg/ml of PYR. (1F) The combination index (CI) vs. fraction-affected (Fa) affected plot was calculated by CompuSyn and depicted the combination effects. Synergistic growth inhibitory effects of pyrotinib (PYR) combined with SHR6390 (SHR) on the EFM-192A cell line. (1G) Proliferation activity of the BT474 cell line was determined by the MTT assay after incubation for 48 h with different concentrations of PYR or SHR. (1H – 1J) The BT474 cell line was treated with PYR or SHR alone or in combination or in sequences (PYR first for 6 h followed by SHR or SHR first for 6 h followed by PYR). (K) The proliferation activity of the BT474 cell line was determined after incubation for 48 h with different concentrations of SHR combination with 0.3, 0.6, 1.2, 2.4, and 4.8 μg/ml of PYR. (1L) The combination index (CI) vs. fraction-affected (Fa) affected plot was calculated by CompuSyn and depicted the combination effects. Synergistic growth inhibitory effects of pyrotinib (PYR) combined with SHR6390 (SHR) on BT474 cell line.
    Figure Legend Snippet: Effects of PYR and SHR on the viability of HER2+/HR+ breast cancer cell lines. (1A) Proliferation activity of the EFM-192A cell line was determined by the MTT assay after incubation for 48 h with different concentrations of PYR or SHR. (1B – 1D) The EFM-192A cell line was treated with PYR or SHR alone or in combination or in sequences (PYR first for 6 h followed by SHR or SHR first for 6 h followed by PYR). (1E) Proliferation activity of the EFM-192A cell line was determined after incubation for 48 h with different concentrations of SHR combination with 0.1875, 0.375, 0.75, 1.5, and 3 μg/ml of PYR. (1F) The combination index (CI) vs. fraction-affected (Fa) affected plot was calculated by CompuSyn and depicted the combination effects. Synergistic growth inhibitory effects of pyrotinib (PYR) combined with SHR6390 (SHR) on the EFM-192A cell line. (1G) Proliferation activity of the BT474 cell line was determined by the MTT assay after incubation for 48 h with different concentrations of PYR or SHR. (1H – 1J) The BT474 cell line was treated with PYR or SHR alone or in combination or in sequences (PYR first for 6 h followed by SHR or SHR first for 6 h followed by PYR). (K) The proliferation activity of the BT474 cell line was determined after incubation for 48 h with different concentrations of SHR combination with 0.3, 0.6, 1.2, 2.4, and 4.8 μg/ml of PYR. (1L) The combination index (CI) vs. fraction-affected (Fa) affected plot was calculated by CompuSyn and depicted the combination effects. Synergistic growth inhibitory effects of pyrotinib (PYR) combined with SHR6390 (SHR) on BT474 cell line.

    Techniques Used: Activity Assay, MTT Assay, Incubation

    23) Product Images from "Anti-Tumor Necrosis Factor Receptor 2 Antibody Combined With Anti-PD-L1 Therapy Exerts Robust Antitumor Effects in Breast Cancer"

    Article Title: Anti-Tumor Necrosis Factor Receptor 2 Antibody Combined With Anti-PD-L1 Therapy Exerts Robust Antitumor Effects in Breast Cancer

    Journal: Frontiers in Cell and Developmental Biology

    doi: 10.3389/fcell.2021.720472

    Tumor necrosis factor receptor 2 is overexpressed in breast tumor and correlated with a poorer overall survival of breast cancer patients. (A) qRT-PCR showed that TNFR2 mRNA level was significantly higher in human breast cancer cell lines (MCF-7, BT549, BT474, and MDA-MB-453) than human mammary epithelial cell MCF-10A (*** p
    Figure Legend Snippet: Tumor necrosis factor receptor 2 is overexpressed in breast tumor and correlated with a poorer overall survival of breast cancer patients. (A) qRT-PCR showed that TNFR2 mRNA level was significantly higher in human breast cancer cell lines (MCF-7, BT549, BT474, and MDA-MB-453) than human mammary epithelial cell MCF-10A (*** p

    Techniques Used: Quantitative RT-PCR, Multiple Displacement Amplification

    24) Product Images from "MCF-7 Drug Resistant Cell Lines Switch Their Lipid Metabolism to Triple Negative Breast Cancer Signature"

    Article Title: MCF-7 Drug Resistant Cell Lines Switch Their Lipid Metabolism to Triple Negative Breast Cancer Signature

    Journal: Cancers

    doi: 10.3390/cancers13235871

    BCCs have a specific lipid signature that differs from each cell line. Principal component analysis (PCA) score plots of the different BCC lines analyzed by LC-MS. Scrutiny of the analyzed BCC lines indicated the similarities and differences between the different lipid species (DG, TG, PC, PE, SM and Cer). It is clear a differential lipid pattern between the studied BCC lines, where the luminal B cell line BT474 and the HER2+ cell line SKBR3 are notably differentiated in their lipid pattern from the rest of cell lines. Moreover, they are notably separated between themselves. Therefore, analyzing the lipid patterns of the BCC lines MCF-7 and its resistant cell lines, MCF-7 Epi R and MCF-7 TAX R , there were significant differences on them. In addition, resistant BCC lines lipid patterns were more similar to the lipid pattern of TNBC line MDA-MB-231 than the lipid profile of sensitive cell line MCF-7 (DG: diacylglycerol, TG: triacylglycerol, PC: phosphatidylcholine, PE: phosphatidylethanolamines, SM: sphingomyelin, Cer: ceramide).
    Figure Legend Snippet: BCCs have a specific lipid signature that differs from each cell line. Principal component analysis (PCA) score plots of the different BCC lines analyzed by LC-MS. Scrutiny of the analyzed BCC lines indicated the similarities and differences between the different lipid species (DG, TG, PC, PE, SM and Cer). It is clear a differential lipid pattern between the studied BCC lines, where the luminal B cell line BT474 and the HER2+ cell line SKBR3 are notably differentiated in their lipid pattern from the rest of cell lines. Moreover, they are notably separated between themselves. Therefore, analyzing the lipid patterns of the BCC lines MCF-7 and its resistant cell lines, MCF-7 Epi R and MCF-7 TAX R , there were significant differences on them. In addition, resistant BCC lines lipid patterns were more similar to the lipid pattern of TNBC line MDA-MB-231 than the lipid profile of sensitive cell line MCF-7 (DG: diacylglycerol, TG: triacylglycerol, PC: phosphatidylcholine, PE: phosphatidylethanolamines, SM: sphingomyelin, Cer: ceramide).

    Techniques Used: Liquid Chromatography with Mass Spectroscopy, Multiple Displacement Amplification

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    ATCC mda mb 231
    Design and functional activity of DARPin-Bn switches. ( A ) Structure model illustrating the interaction between the HER2 extracellular domain (ECD) and DARPin-Bn switches G3-Bn and 9.29-Bn. HER2 ECD (solid surface, PDB ID: 1N8Z), G3-Bn (yellow ribbon, PDB ID: 4HRN), and 9.29-Bn (red ribbon, PDB ID: 4HRL) were aligned by HER2 subdomains (colored individually). DARPins-Bn were constructed de novo, and the free energy was minimized in the elongated conformation, barnase (cyan ribbon, PDB ID: 1BRS). ( B and С ) DARPins-Bn specifically bind to the surface of HER2 + cells. ( B ) Confocal scanning microscopy. Bright field, Top Panels ; FITC (λ ex = 488 nm, λ em = 492 to 550 nm) and Hoechst 33342 (λ ex = 405 nm, λ em = 410 to 520 nm) overlay, Bottom Panels . ( C ) Flow cytometry. The histograms show the fluorescence distribution of unlabeled (autofluorescence) and labeled cells. HER2 + BT-474 cells, <t>MDA-MB-231</t> cells with a low level of HER2 expression, and control HER2 − CHO cells were stained with 9.29-Bn and G3-Bn labeled with FITC. Trastuzumab was used as a positive control. ( D ) DARPin-Bn switches mediate specific cytotoxicity after a prolonged incubation with HER2 + cells. Resazurin cell viability assay of BT-474, MDA-MB-231, and CHO cells treated with 9.29-Bn, G3-Bn, and free barnase after 10 d of incubation. Data are represented as means ± SDs.
    Mda Mb 231, supplied by ATCC, used in various techniques. Bioz Stars score: 88/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    bt 474  (ATCC)
    98
    ATCC bt 474
    The g3mclass autoclassification on three genes. ( A ) The g3mclass -selected models. The 5-class model parameters for ESR1 mRNA: class − 1 (0.49 ± 0.57; weight 0.25); class 0 (3.34 ± 3.18; weight 0.27), class 1 (17.4 ± 9.22; weight 0.25), class 2 (63.4 ± 29.2; weight 0.19), class 3 (99.5 ± 105; weight 0.05). The 4-class model parameters for PGR mRNA: class − 1 (0.30 ± 0.43; weight 0.48); class 0 (3.09 ± 2.77; weight 0.26), class 1 (7.88 ± 3.49; weight 0.13), class 2 (63.4 ± 29.2; weight 0.12). The 4-class model parameters for ERBB2 mRNA as above in Fig. 2 . ( B )–( E ) The g3mclass -created classification heatmaps. The s.cutoff classification heatmaps on ESR1, PGR, and ERBB2 for mammoplasties ( B ), IBC ( C ), DCIS ( D ), and IBC cell lines ( E ). The IHC score for ER-positive (1), and ER-negative (0) cancers are on the top of heatmaps ( C , D ). In E, human breast cancer cell lines: MDA-MB-231 (triple-negative), SK-BR-3 (HER2 overexpressing), <t>BT-474</t> (luminal B), T47D (luminal A), MCF7 (luminal A). Bar, heatmap’s color scale for classes.
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    90
    ATCC bt474 her2 cells
    Bar graphs representing [ 68 Ga]P5 and [ 68 Ga]P6 binding to <t>BT474</t> <t>(HER2+)</t> and MDA-MB-231 (HER2−) <t>cells.</t> ( A ) [ 68 Ga]P5 had significantly higher % binding per mg of protein to BT474 cells (0.68 ± 0.20) versus MDA-MB-231 cells (0.47 ± 0.05) after 15 min of incubation. ( n = 6) ( B ) [ 68 Ga]P6 also had significantly higher binding to BT474 cells (0.55 ± 0.21) versus MDA-MB-231 cells (0.34 ± 0.12) after 30 min. ( n = 12) * p
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    Design and functional activity of DARPin-Bn switches. ( A ) Structure model illustrating the interaction between the HER2 extracellular domain (ECD) and DARPin-Bn switches G3-Bn and 9.29-Bn. HER2 ECD (solid surface, PDB ID: 1N8Z), G3-Bn (yellow ribbon, PDB ID: 4HRN), and 9.29-Bn (red ribbon, PDB ID: 4HRL) were aligned by HER2 subdomains (colored individually). DARPins-Bn were constructed de novo, and the free energy was minimized in the elongated conformation, barnase (cyan ribbon, PDB ID: 1BRS). ( B and С ) DARPins-Bn specifically bind to the surface of HER2 + cells. ( B ) Confocal scanning microscopy. Bright field, Top Panels ; FITC (λ ex = 488 nm, λ em = 492 to 550 nm) and Hoechst 33342 (λ ex = 405 nm, λ em = 410 to 520 nm) overlay, Bottom Panels . ( C ) Flow cytometry. The histograms show the fluorescence distribution of unlabeled (autofluorescence) and labeled cells. HER2 + BT-474 cells, MDA-MB-231 cells with a low level of HER2 expression, and control HER2 − CHO cells were stained with 9.29-Bn and G3-Bn labeled with FITC. Trastuzumab was used as a positive control. ( D ) DARPin-Bn switches mediate specific cytotoxicity after a prolonged incubation with HER2 + cells. Resazurin cell viability assay of BT-474, MDA-MB-231, and CHO cells treated with 9.29-Bn, G3-Bn, and free barnase after 10 d of incubation. Data are represented as means ± SDs.

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Switchable targeting of solid tumors by BsCAR T cells

    doi: 10.1073/pnas.2210562119

    Figure Lengend Snippet: Design and functional activity of DARPin-Bn switches. ( A ) Structure model illustrating the interaction between the HER2 extracellular domain (ECD) and DARPin-Bn switches G3-Bn and 9.29-Bn. HER2 ECD (solid surface, PDB ID: 1N8Z), G3-Bn (yellow ribbon, PDB ID: 4HRN), and 9.29-Bn (red ribbon, PDB ID: 4HRL) were aligned by HER2 subdomains (colored individually). DARPins-Bn were constructed de novo, and the free energy was minimized in the elongated conformation, barnase (cyan ribbon, PDB ID: 1BRS). ( B and С ) DARPins-Bn specifically bind to the surface of HER2 + cells. ( B ) Confocal scanning microscopy. Bright field, Top Panels ; FITC (λ ex = 488 nm, λ em = 492 to 550 nm) and Hoechst 33342 (λ ex = 405 nm, λ em = 410 to 520 nm) overlay, Bottom Panels . ( C ) Flow cytometry. The histograms show the fluorescence distribution of unlabeled (autofluorescence) and labeled cells. HER2 + BT-474 cells, MDA-MB-231 cells with a low level of HER2 expression, and control HER2 − CHO cells were stained with 9.29-Bn and G3-Bn labeled with FITC. Trastuzumab was used as a positive control. ( D ) DARPin-Bn switches mediate specific cytotoxicity after a prolonged incubation with HER2 + cells. Resazurin cell viability assay of BT-474, MDA-MB-231, and CHO cells treated with 9.29-Bn, G3-Bn, and free barnase after 10 d of incubation. Data are represented as means ± SDs.

    Article Snippet: Cell lines of human ductal carcinoma BT-474 (HTB-2; ATCC), MDA-MB-231 (HTB-26; ATCC), HEK293T lentiviral packaging cell line (Clontech), and CHO (Russian Cell Culture Collection) were incubated in a humidified atmosphere with 5% CO2 at 37 °C.

    Techniques: Functional Assay, Activity Assay, Construct, Microscopy, Flow Cytometry, Fluorescence, Labeling, Multiple Displacement Amplification, Expressing, Staining, Positive Control, Incubation, Viability Assay

    The g3mclass autoclassification on three genes. ( A ) The g3mclass -selected models. The 5-class model parameters for ESR1 mRNA: class − 1 (0.49 ± 0.57; weight 0.25); class 0 (3.34 ± 3.18; weight 0.27), class 1 (17.4 ± 9.22; weight 0.25), class 2 (63.4 ± 29.2; weight 0.19), class 3 (99.5 ± 105; weight 0.05). The 4-class model parameters for PGR mRNA: class − 1 (0.30 ± 0.43; weight 0.48); class 0 (3.09 ± 2.77; weight 0.26), class 1 (7.88 ± 3.49; weight 0.13), class 2 (63.4 ± 29.2; weight 0.12). The 4-class model parameters for ERBB2 mRNA as above in Fig. 2 . ( B )–( E ) The g3mclass -created classification heatmaps. The s.cutoff classification heatmaps on ESR1, PGR, and ERBB2 for mammoplasties ( B ), IBC ( C ), DCIS ( D ), and IBC cell lines ( E ). The IHC score for ER-positive (1), and ER-negative (0) cancers are on the top of heatmaps ( C , D ). In E, human breast cancer cell lines: MDA-MB-231 (triple-negative), SK-BR-3 (HER2 overexpressing), BT-474 (luminal B), T47D (luminal A), MCF7 (luminal A). Bar, heatmap’s color scale for classes.

    Journal: Scientific Reports

    Article Title: The g3mclass is a practical software for multiclass classification on biomarkers

    doi: 10.1038/s41598-022-23438-9

    Figure Lengend Snippet: The g3mclass autoclassification on three genes. ( A ) The g3mclass -selected models. The 5-class model parameters for ESR1 mRNA: class − 1 (0.49 ± 0.57; weight 0.25); class 0 (3.34 ± 3.18; weight 0.27), class 1 (17.4 ± 9.22; weight 0.25), class 2 (63.4 ± 29.2; weight 0.19), class 3 (99.5 ± 105; weight 0.05). The 4-class model parameters for PGR mRNA: class − 1 (0.30 ± 0.43; weight 0.48); class 0 (3.09 ± 2.77; weight 0.26), class 1 (7.88 ± 3.49; weight 0.13), class 2 (63.4 ± 29.2; weight 0.12). The 4-class model parameters for ERBB2 mRNA as above in Fig. 2 . ( B )–( E ) The g3mclass -created classification heatmaps. The s.cutoff classification heatmaps on ESR1, PGR, and ERBB2 for mammoplasties ( B ), IBC ( C ), DCIS ( D ), and IBC cell lines ( E ). The IHC score for ER-positive (1), and ER-negative (0) cancers are on the top of heatmaps ( C , D ). In E, human breast cancer cell lines: MDA-MB-231 (triple-negative), SK-BR-3 (HER2 overexpressing), BT-474 (luminal B), T47D (luminal A), MCF7 (luminal A). Bar, heatmap’s color scale for classes.

    Article Snippet: The cell lines MCF-7T-47D, MDA-MB-231, SK-BR-3, and BT-474 were purchased from the American Type Culture Collection and cultured accordingly.

    Techniques: Immunohistochemistry, Multiple Displacement Amplification

    LINC00847-FOXA1 and CTD-2566J3.1 GINS2 are essential genes for proliferation and survival. ( A ) DEMETER score values for each mRNA reported in the co-expressed mRNAs/lncRNAs pairs of 3D cultures and TCGA data. ( B , C ) The DEMETER score is shown against the gene expression of FOXA1 and GINS2 ; the stars represent BT-474 cell line and the color dots represent the breast cancer subtype of each cell line.

    Journal: Cells

    Article Title: Breast Cancer Cells Reprogram the Oncogenic lncRNAs/mRNAs Coexpression Networks in Three-Dimensional Microenvironment

    doi: 10.3390/cells11213458

    Figure Lengend Snippet: LINC00847-FOXA1 and CTD-2566J3.1 GINS2 are essential genes for proliferation and survival. ( A ) DEMETER score values for each mRNA reported in the co-expressed mRNAs/lncRNAs pairs of 3D cultures and TCGA data. ( B , C ) The DEMETER score is shown against the gene expression of FOXA1 and GINS2 ; the stars represent BT-474 cell line and the color dots represent the breast cancer subtype of each cell line.

    Article Snippet: BT-474 cell lines were obtained from ATCC (Manassas, Virginia) and cultured in Hybri-Care medium and 10% fetal bovine serum (FBS).

    Techniques: Expressing

    Co-expression lncRNAs/mRNAs networks common in BT-474 cell grown in 3D cultures and breast tumors (TCGA). The diagram shows how the cellular processes associated to cancer hallmarks, could be regulated by ncRNAs/mRNAs pairs that are common in BT-474 cultures and breast biopsies. Blue dot: lncRNAs; green dot: mRNAs; pink dot; multiple colors; cancer hallmarks and therapy resistance.

    Journal: Cells

    Article Title: Breast Cancer Cells Reprogram the Oncogenic lncRNAs/mRNAs Coexpression Networks in Three-Dimensional Microenvironment

    doi: 10.3390/cells11213458

    Figure Lengend Snippet: Co-expression lncRNAs/mRNAs networks common in BT-474 cell grown in 3D cultures and breast tumors (TCGA). The diagram shows how the cellular processes associated to cancer hallmarks, could be regulated by ncRNAs/mRNAs pairs that are common in BT-474 cultures and breast biopsies. Blue dot: lncRNAs; green dot: mRNAs; pink dot; multiple colors; cancer hallmarks and therapy resistance.

    Article Snippet: BT-474 cell lines were obtained from ATCC (Manassas, Virginia) and cultured in Hybri-Care medium and 10% fetal bovine serum (FBS).

    Techniques: Expressing

    Generation and morphology of 3D organotypic cultures. ( A ) Growth kinetics of BT-474 cancer cells grown in 2D monolayers and 3D cell cultures observed in optical microscopy (20× and 40×) during 0–6 days incubation over Geltrex. ( B ) Quantification of the diameter of round-like structures in 3D cell cultures. ( C ) Quantification of the number of structures in 3D cultures at optimized seeding densities of 25,000 cells/well plate. ( D ) Hematoxylin and eosin staining of BT-474 cancer cells grown in 2D and 3D culture conditions. ( E ) Color image of 2D monolayers and 3D cell cultures using Identify Primary Objects module. Filter Objects module discards objects that have irregular shape or cell debris (magenta lines). ( F ) Quantification of number of nuclei present in 3D round-like structures. Values represent the mean ± SD from 3 independent experiments, using an unpaired Student’s t -test with unequal variances; * p

    Journal: Cells

    Article Title: Breast Cancer Cells Reprogram the Oncogenic lncRNAs/mRNAs Coexpression Networks in Three-Dimensional Microenvironment

    doi: 10.3390/cells11213458

    Figure Lengend Snippet: Generation and morphology of 3D organotypic cultures. ( A ) Growth kinetics of BT-474 cancer cells grown in 2D monolayers and 3D cell cultures observed in optical microscopy (20× and 40×) during 0–6 days incubation over Geltrex. ( B ) Quantification of the diameter of round-like structures in 3D cell cultures. ( C ) Quantification of the number of structures in 3D cultures at optimized seeding densities of 25,000 cells/well plate. ( D ) Hematoxylin and eosin staining of BT-474 cancer cells grown in 2D and 3D culture conditions. ( E ) Color image of 2D monolayers and 3D cell cultures using Identify Primary Objects module. Filter Objects module discards objects that have irregular shape or cell debris (magenta lines). ( F ) Quantification of number of nuclei present in 3D round-like structures. Values represent the mean ± SD from 3 independent experiments, using an unpaired Student’s t -test with unequal variances; * p

    Article Snippet: BT-474 cell lines were obtained from ATCC (Manassas, Virginia) and cultured in Hybri-Care medium and 10% fetal bovine serum (FBS).

    Techniques: Microscopy, Incubation, Staining

    Bar graphs representing [ 68 Ga]P5 and [ 68 Ga]P6 binding to BT474 (HER2+) and MDA-MB-231 (HER2−) cells. ( A ) [ 68 Ga]P5 had significantly higher % binding per mg of protein to BT474 cells (0.68 ± 0.20) versus MDA-MB-231 cells (0.47 ± 0.05) after 15 min of incubation. ( n = 6) ( B ) [ 68 Ga]P6 also had significantly higher binding to BT474 cells (0.55 ± 0.21) versus MDA-MB-231 cells (0.34 ± 0.12) after 30 min. ( n = 12) * p

    Journal: Diagnostics

    Article Title: Evaluation of 68Ga-Radiolabeled Peptides for HER2 PET Imaging

    doi: 10.3390/diagnostics12112710

    Figure Lengend Snippet: Bar graphs representing [ 68 Ga]P5 and [ 68 Ga]P6 binding to BT474 (HER2+) and MDA-MB-231 (HER2−) cells. ( A ) [ 68 Ga]P5 had significantly higher % binding per mg of protein to BT474 cells (0.68 ± 0.20) versus MDA-MB-231 cells (0.47 ± 0.05) after 15 min of incubation. ( n = 6) ( B ) [ 68 Ga]P6 also had significantly higher binding to BT474 cells (0.55 ± 0.21) versus MDA-MB-231 cells (0.34 ± 0.12) after 30 min. ( n = 12) * p

    Article Snippet: MDA-MB-231 (HER2−) and BT474 (HER2+) cells were purchased from American Type Culture Collection (Manassas, VA, USA).

    Techniques: Binding Assay, Multiple Displacement Amplification, Incubation

    One hour post-injection biodistribution for [ 68 Ga]P6 in athymic nude mice. [ 68 Ga]P6 showed significantly higher binding to HER2-positive tumors ( n = 4) versus HER2-negative tumors ( n = 3). * p

    Journal: Diagnostics

    Article Title: Evaluation of 68Ga-Radiolabeled Peptides for HER2 PET Imaging

    doi: 10.3390/diagnostics12112710

    Figure Lengend Snippet: One hour post-injection biodistribution for [ 68 Ga]P6 in athymic nude mice. [ 68 Ga]P6 showed significantly higher binding to HER2-positive tumors ( n = 4) versus HER2-negative tumors ( n = 3). * p

    Article Snippet: MDA-MB-231 (HER2−) and BT474 (HER2+) cells were purchased from American Type Culture Collection (Manassas, VA, USA).

    Techniques: Injection, Mouse Assay, Binding Assay

    PET images of the 50-60 min frame with a SUV scale of 0.2-3. ( A ) [ 68 Ga]P6 showed stronger binding to the HER2-positive tumor with an SUV mean value of 0.72 ± 0.16, while ( B ) less binding was seen in the HER2-negative tumor shown by the lower SUV mean value of 0.61 ± 0.19.

    Journal: Diagnostics

    Article Title: Evaluation of 68Ga-Radiolabeled Peptides for HER2 PET Imaging

    doi: 10.3390/diagnostics12112710

    Figure Lengend Snippet: PET images of the 50-60 min frame with a SUV scale of 0.2-3. ( A ) [ 68 Ga]P6 showed stronger binding to the HER2-positive tumor with an SUV mean value of 0.72 ± 0.16, while ( B ) less binding was seen in the HER2-negative tumor shown by the lower SUV mean value of 0.61 ± 0.19.

    Article Snippet: MDA-MB-231 (HER2−) and BT474 (HER2+) cells were purchased from American Type Culture Collection (Manassas, VA, USA).

    Techniques: Positron Emission Tomography, Binding Assay

    Biodistribution of [ 68 Ga]P5 two hours post-injection in athymic nude mice bearing HER2-negative (MDA-MB-231 cells) and HER2-positive tumors (BT474 cells). Each organ was placed on the HIDEX AMG to determine the amount of [ 68 Ga]P5 uptake. HER2-positive tumors showed significantly higher binding than the HER2-negative tumors ( n = 4). * p

    Journal: Diagnostics

    Article Title: Evaluation of 68Ga-Radiolabeled Peptides for HER2 PET Imaging

    doi: 10.3390/diagnostics12112710

    Figure Lengend Snippet: Biodistribution of [ 68 Ga]P5 two hours post-injection in athymic nude mice bearing HER2-negative (MDA-MB-231 cells) and HER2-positive tumors (BT474 cells). Each organ was placed on the HIDEX AMG to determine the amount of [ 68 Ga]P5 uptake. HER2-positive tumors showed significantly higher binding than the HER2-negative tumors ( n = 4). * p

    Article Snippet: MDA-MB-231 (HER2−) and BT474 (HER2+) cells were purchased from American Type Culture Collection (Manassas, VA, USA).

    Techniques: Injection, Mouse Assay, Multiple Displacement Amplification, Binding Assay