Journal: Journal of Cancer

Article Title: Tumor Suppressor RARRES1 Regulates DLG2, PP2A, VCP, EB1, and Ankrd26

doi:

Figure Lengend Snippet: Sample mixture loaded on each gel : Each sample was composed of equal amounts of RARRES1 knock-down PWR-1E and control lysates. Cy3 and Cy5 dyes were equally swapped among RARES1 knock-down and control samples. The internal standard was labeled with Cy2 and consisted of 12.5 µg of each knock-down and control. The sample loaded on the pick-gel was composed of 300 µg of each sample and control.

Article Snippet: Western Blotting was performed as described previously using the following antibodies: Rabbit monoclonal to PP2A (cat. ab32141, Abcam, Cambridge, MA); Rabbit polyclonal to PSD93 (cat. ab2930, Abcam, Cambridge, MA); Goat polyclonal to RARRES1 (R&D systems, Mineapolis, MN); Mouse monoclonal to VCP (cat. ab11433, Abcam, Cambridge, MA).

Techniques: Knockdown, Control, Labeling

Journal: Journal of Cancer

Article Title: Tumor Suppressor RARRES1 Regulates DLG2, PP2A, VCP, EB1, and Ankrd26

doi:

Figure Lengend Snippet: DIGE data : Representation set of one of the 4 biological replicates. 25 µg of control PWR-1E cells lysates labeled with 200 pmoles of Cy3 (green); 25 µg of RARRES1 knock-down PWR-1E cells labeled with 200 pmoles of Cy5 (red) and a normalization pool composed of a 50:50 mixture of sample and control labeled with Cy2 (blue). The first dimension separation was performed on a 24 cm non-linear IPG strips and the second dimension separation was accomplished on a 5-20% gradient polyacrylamide gel with 2 built-in spot picking references. Gels were scanned at a 100 µm resolution using excitation and emission wavelength that corresponds to each dye. Combined Cy2, Cy3, and Cy5 signals are represented in the top gel.

Article Snippet: Western Blotting was performed as described previously using the following antibodies: Rabbit monoclonal to PP2A (cat. ab32141, Abcam, Cambridge, MA); Rabbit polyclonal to PSD93 (cat. ab2930, Abcam, Cambridge, MA); Goat polyclonal to RARRES1 (R&D systems, Mineapolis, MN); Mouse monoclonal to VCP (cat. ab11433, Abcam, Cambridge, MA).

Techniques: Control, Labeling, Knockdown

Journal: Journal of Cancer

Article Title: Tumor Suppressor RARRES1 Regulates DLG2, PP2A, VCP, EB1, and Ankrd26

doi:

Figure Lengend Snippet: Sypro Ruby Pick gel : loaded with 300µg of PWR-1E RARRES1 knock-down protein extracts and 300µg of control sample. The green dots represent the proteins that were detected by the Decyder software. Annotated proteins are those that were picked for mass spectrometry analysis. These spots were identified as differentially-expressed across the 4 biological replicates in the analytical gels. The coordinates of these spots were recorded, matched, and excised from the Pick gel for identification.

Article Snippet: Western Blotting was performed as described previously using the following antibodies: Rabbit monoclonal to PP2A (cat. ab32141, Abcam, Cambridge, MA); Rabbit polyclonal to PSD93 (cat. ab2930, Abcam, Cambridge, MA); Goat polyclonal to RARRES1 (R&D systems, Mineapolis, MN); Mouse monoclonal to VCP (cat. ab11433, Abcam, Cambridge, MA).

Techniques: Knockdown, Control, Software, Mass Spectrometry

Journal: Journal of Cancer

Article Title: Tumor Suppressor RARRES1 Regulates DLG2, PP2A, VCP, EB1, and Ankrd26

doi:

Figure Lengend Snippet: Differentially expressed proteins : a) 2- Cropped DIGE; b) three-dimensional view, and c) logarithmic representation of the 4 four biological replicates of Dlg-2 upregulation, PP2A and VCP down-regulation as a result of RARRES1 knock-down in PWR-1E cells.

Article Snippet: Western Blotting was performed as described previously using the following antibodies: Rabbit monoclonal to PP2A (cat. ab32141, Abcam, Cambridge, MA); Rabbit polyclonal to PSD93 (cat. ab2930, Abcam, Cambridge, MA); Goat polyclonal to RARRES1 (R&D systems, Mineapolis, MN); Mouse monoclonal to VCP (cat. ab11433, Abcam, Cambridge, MA).

Techniques: Knockdown

Journal: Journal of Cancer

Article Title: Tumor Suppressor RARRES1 Regulates DLG2, PP2A, VCP, EB1, and Ankrd26

doi:

Figure Lengend Snippet: Western Blot Validation : of a) RARRES1 knock-down, b) Dlg2 homologue, c) PP2A, d) VCP and e) GAPDH loading control. Lane 1 scrambled siRNA nucleofected PWR-1E lysates; lane 2 RARRES1 siRNA nucleofected PWR-1E lysates.

Article Snippet: Western Blotting was performed as described previously using the following antibodies: Rabbit monoclonal to PP2A (cat. ab32141, Abcam, Cambridge, MA); Rabbit polyclonal to PSD93 (cat. ab2930, Abcam, Cambridge, MA); Goat polyclonal to RARRES1 (R&D systems, Mineapolis, MN); Mouse monoclonal to VCP (cat. ab11433, Abcam, Cambridge, MA).

Techniques: Western Blot, Biomarker Discovery, Knockdown, Control

Journal: Biology of Reproduction

Article Title: Random Allocation of Blastomere Descendants to the Trophectoderm and ICM of the Bovine Blastocyst 1

doi: 10.1095/biolreprod.116.141200

Figure Lengend Snippet: Immunodetection of SOX2. Immunodetection of Sox2 (green) and labeling of DNA by DAPI (blue) was performed in each group of bovine blastocysts (bar = 50 μm). The total number of DAPI- and SOX2-positive cells was determined on z-projections using the cell counter plugin of ImageJ software.

Article Snippet: The samples were rinsed in PBS and then incubated with 2% (w/v) BSA-PBS for 1 h. Then the embryos were incubated in mouse monoclonal anti-Sox2 antibody (1:50 in 2% BSA-PBS; R&D Systems) for 4 h at room temperature.

Techniques: Immunodetection, Labeling, Software

Journal: Heliyon

Article Title: Novel roles for HMGA2 isoforms in regulating oxidative stress and sensitizing to RSL3-Induced ferroptosis in prostate cancer cells

doi: 10.1016/j.heliyon.2023.e14810

Figure Lengend Snippet: HMGA2-WT and -TR Expression in PCa and Breast cancer Cell Lines and Patients Tissue (A) Western blot analysis shows the expression of HMGA2 protein in prostate cancer and breast cancer cell lines, HMGA2-WT displays a higher molecular weight at about 23 KDa while the HMGA2-TR displays a lower molecular of about 17 KDa. (B) Western blot analysis measured the protein levels of both HMGA2-WT and -TR in prostate cancer patient tissue displaying different Stage/Gleason scores. (C) RT-PCR measured the mRNA levels in cDNA array. (D) Immunohistochemical analysis was performed using prostate tissue microarray with HMGA2 antibody in normal prostate tissue, hyperplasia, and chronic inflammation prostate tissue. Scale bar; 100 μm. Magnification 20X, inset 40X. Statistical analysis was conducted utilizing GraphPad Prism; (**** p < 0.0001, ***p < 0.001, **p < 0.01). Bars represents SD of the mean. Alpha-tubulin was used as a loading control. Results are representative of 3 independent experiments.

Article Snippet: Goat monoclonal anti -HMGA2 was purchased from R&D Systems (Minneapolis, MN) for immunofluorescence and anti -HMGA2 (ThermoFisher, Waltham, MA) for Western blot analyses.

Techniques: Expressing, Western Blot, Molecular Weight, Reverse Transcription Polymerase Chain Reaction, Immunohistochemical staining, Microarray, Control

Journal: Heliyon

Article Title: Novel roles for HMGA2 isoforms in regulating oxidative stress and sensitizing to RSL3-Induced ferroptosis in prostate cancer cells

doi: 10.1016/j.heliyon.2023.e14810

Figure Lengend Snippet: LNCaP Cells Stably Overexpressing Truncated HMGA2 Display Cytoplasmic Localization and Increased Reactive Oxygen Species LNCaP cells stably overexpressing wild-type HMGA2 (HMGA2-WT), truncated HMGA2 (HMGA2-TR) or empty vector control (Neo) were assayed by (A) immunofluorescence with anti -HMGA2 antibody, and DAPI to stain the nucleus. HMGA2-WT displays a nuclear/cytoplasmic localization while HMGA2-TR is predominantly cytoplasmic. Magnification 40X. (B) Overexpression of HMGA2-WT and HMGA2-TR is associated with morphological changes compared to Neo control as shown by brightfield microscopy. Magnification 20X; Inset 40X. (C) Western blot analysis shows expression of HMGA2 in LNCaP Neo, LNCaP HMGA2-WT, and LNCaP HMGA2-TR with quantification of the blot using Image J (NIH). (D) ROS was measured using DCFDA which shows that HMGA2-TR increases ROS generation. Metabolomics analysis reveals that (E) NADP/NADPH ratio is increased while (F) there is significant decrease in GSH/GSSG ratio in HMGA2-TR compared to HMGA2-WT and Neo. Statistical analysis was done with GraphPad Prism; (**** p < 0.0001, ***p < 0.001, **p < 0.01, * p < 0.05). Bars represents SD of the mean. Results are representative of 3 independent experiments.

Article Snippet: Goat monoclonal anti -HMGA2 was purchased from R&D Systems (Minneapolis, MN) for immunofluorescence and anti -HMGA2 (ThermoFisher, Waltham, MA) for Western blot analyses.

Techniques: Stable Transfection, Plasmid Preparation, Control, Immunofluorescence, Staining, Over Expression, Microscopy, Western Blot, Expressing

Journal: Heliyon

Article Title: Novel roles for HMGA2 isoforms in regulating oxidative stress and sensitizing to RSL3-Induced ferroptosis in prostate cancer cells

doi: 10.1016/j.heliyon.2023.e14810

Figure Lengend Snippet: RNA-Seq Reveals Upregulated and Downregulated Genes in HMGA2-WT and HMGA2-TR Cells mRNA from LNCaP Neo, HMGA2-WT and HMGA2-TR cells was utilized to perform RNA-Seq analysis. Schematic view of data from RNA-seq from HMGA2-WT and HMGA2-TR (A) upregulated, and (B) down-regulated genes as compared to Neo. (C) Further analysis was performed using GOrilla software which revealed upregulated (red) and downregulated (green) genes. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: Goat monoclonal anti -HMGA2 was purchased from R&D Systems (Minneapolis, MN) for immunofluorescence and anti -HMGA2 (ThermoFisher, Waltham, MA) for Western blot analyses.

Techniques: RNA Sequencing, Software

Journal: Heliyon

Article Title: Novel roles for HMGA2 isoforms in regulating oxidative stress and sensitizing to RSL3-Induced ferroptosis in prostate cancer cells

doi: 10.1016/j.heliyon.2023.e14810

Figure Lengend Snippet: RNA-Seq Reveals that Truncated HMGA2 is Involved in Oxidative Stress Gene Set Enrichment Analysis (GSEA) of RNA-Seq data detects pathways regulated by (A) HMGA2-TR and (B) HMGA2-WT cells when compared to Neo control. Ingenuity Pathway Analysis (IPA) shows prediction of upregulated (orange) and downregulated (blue) pathways in (C) HMGA2-TR and (D) HMGA2-WT cells compared to Neo control. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: Goat monoclonal anti -HMGA2 was purchased from R&D Systems (Minneapolis, MN) for immunofluorescence and anti -HMGA2 (ThermoFisher, Waltham, MA) for Western blot analyses.

Techniques: RNA Sequencing, Control

Journal: Heliyon

Article Title: Novel roles for HMGA2 isoforms in regulating oxidative stress and sensitizing to RSL3-Induced ferroptosis in prostate cancer cells

doi: 10.1016/j.heliyon.2023.e14810

Figure Lengend Snippet: G3BP1 Interacts with cytoplasmic HMGA2 (A) Nuclear fraction of HMGA2-WT cells and whole cell lysates of HMGA2-TR cells were utilized to perform immunoprecipitation (IP) with HMGA2 antibody or IgG control followed by proteomics to identify HMGA2-interacting proteins. Venn diagram represents different proteins that interact with HMGA2, but not IgG, in HMGA2-WT and HMGA2-TR cells with KRT1 common to both. (B) Immunofluorescence shows the expression of G3BP1 in LNCaP Neo, HMGA2-WT and HMGA2-TR cells. (C) We isolated nuclear fraction or whole cell lysate from HMGA2-WT cells or whole cell lysate from HMGA2-TR cells and performed IP with HMGA2 antibody or IgG control followed by Western blot with G3BP1. We performed transient knockdown of G3BP1 in HMGA2-WT and HMGA2-TR cells followed by (D) immunofluorescence with HMGA2 and G3BP1 antibody including DAPI stain for the nucleus, or (E) ROS analysis using DCFDA stain. Statistical analysis was done with GraphPad Prism; (**** p < 0.0001, * p < 0.05). Bars represents SD of the mean. Magnification 40X. Results are representative of 3 independent experiments.

Article Snippet: Goat monoclonal anti -HMGA2 was purchased from R&D Systems (Minneapolis, MN) for immunofluorescence and anti -HMGA2 (ThermoFisher, Waltham, MA) for Western blot analyses.

Techniques: Immunoprecipitation, Control, Immunofluorescence, Expressing, Isolation, Western Blot, Knockdown, Staining

Journal: Heliyon

Article Title: Novel roles for HMGA2 isoforms in regulating oxidative stress and sensitizing to RSL3-Induced ferroptosis in prostate cancer cells

doi: 10.1016/j.heliyon.2023.e14810

Figure Lengend Snippet: Truncated HMGA2 Promotes Sensitivity to Ferroptosis Induction that is Exacerbated by G3BP1 Silencing (A) LNCaP Neo, HMGA2-WT cells or HMGA2-TR cells were treated with various concentrations of ferroptosis inducer RSL3 for 72 h followed by cell viability analysis using MTS assay. RSL3 treatments were repeated in the presence or absence of 1 μM ferrostatin-1 and cell viability assayed in (B) HMGA2-WT cells, or (C) HMGA2-TR cells. (D) HMGA2-TR cells were treated with control or G3BP1 siRNA followed by Western blot analysis. (E) HMGA2-TR cells were co-treated with control or G3BP1 siRNA plus 4 μM RSL3 followed by MTS assay. Statistical analysis was done with GraphPad Prism; (**** p < 0.0001, *** p < 0.001, **p < 0.01, * p < 0.05). Bars represents SD of the mean. Results are representative of 3 independent experiments.

Article Snippet: Goat monoclonal anti -HMGA2 was purchased from R&D Systems (Minneapolis, MN) for immunofluorescence and anti -HMGA2 (ThermoFisher, Waltham, MA) for Western blot analyses.

Techniques: MTS Assay, Control, Western Blot

Journal: Heliyon

Article Title: Novel roles for HMGA2 isoforms in regulating oxidative stress and sensitizing to RSL3-Induced ferroptosis in prostate cancer cells

doi: 10.1016/j.heliyon.2023.e14810

Figure Lengend Snippet: Endogenous HMGA2 and G3BP1 Mediate Ferroptosis in PC3 Cells (A) PC3 PCa cells were treated with control or HMGA2 siRNA plus or minus 4 μM RSL3 and 1 μM ferrostatin-1 for 72 h followed by cell viability analysis using MTS assay. (B) Knockdown of HMGA2 was confirmed by Western blot analysis. (C) PC3 cells were treated with control or G3BP1 siRNA plus or minus 4 μM RSL3 and 1 μM ferrostatin-1 for 72 h followed by cell viability analysis using MTS assay. (B) Knockdown of G3BP1 was confirmed by Western blot analysis. Statistical analysis was done with GraphPad Prism; (**** p < 0.0001, *** p < 0.001, **p < 0.01, * p < 0.05). Bars represents SD of the mean. Results are representative of 3 independent experiments.

Article Snippet: Goat monoclonal anti -HMGA2 was purchased from R&D Systems (Minneapolis, MN) for immunofluorescence and anti -HMGA2 (ThermoFisher, Waltham, MA) for Western blot analyses.

Techniques: Control, MTS Assay, Knockdown, Western Blot

Journal: bioRxiv

Article Title: A Novel Mouse Model Reveals a Role for Mitochondria in Early Lineage Specification and Gastrulation

doi: 10.1101/2025.07.14.664670

Figure Lengend Snippet: A. Immunofluorescence staining of Brachyury (magenta), SOX2 (green), and nuclei (DAPI, gray) in sagittal sections of E7.5 embryos. In control ( R26R-mitoAtDarT ) embryos, SOX2 is expressed in the epiblast and upregulated in neural ectoderm progenitors and the chorion, while Brachyury expression is appropriately restricted to mesodermal cells within the notochord region. In R26-mitoAtDarT embryos, Brachyury expression is aberrantly extended into extraembryonic regions, indicating disrupted mesoderm patterning during gastrulation. SOX2 expression remains intact. Scale bars, 20 μm. N = 5 embryos per genotype. B. Coronal sections of E7.5 embryos stained for SOX17 (magenta) and nuclei (DAPI, gray). In control embryos, SOX17 expression is localized to the visceral endoderm with sparse positive cells in the allantois. In R26-mitoAtDarT embryos, SOX17-positive cells surround the embryo as expected but show increased numbers and aggregation in the allantoic bud region. Scale bars, 20 μm. N = 3 embryos per genotype.

Article Snippet: Cryosections of post-implantation embryos were washed with 0.3% PBST three times for 20 minutes each to ensure permeabilization and complete removal of the OCT. Tissues were then blocked in 10% heat-inactivated goat serum (Himedia) and 1% BSA (Sigma) in 0.3 % PBST for 1 hour, followed by overnight incubation with primary antibodies: CdX2 (Biogenix-MU392A), SOX2 (Invitrogen-14-9811-82), SOX17 (R&D systems-af1924), Brachyury (Abcam-EPR18113), TOMM20 (Thermofisher-MA532148), FLAG-tag (Sigma-F1804), phospho-Histone H3 (pHH3) (Cell Signalling Technology - 9706S), and Cleaved caspase (Cell Signalling Technology - 9661S) at a 1:200 concentration diluted in the blocking buffer, overnight at 4°C.

Techniques: Immunofluorescence, Staining, Control, Expressing

Journal: bioRxiv

Article Title: A Novel Mouse Model Reveals a Role for Mitochondria in Early Lineage Specification and Gastrulation

doi: 10.1101/2025.07.14.664670

Figure Lengend Snippet: A. E3.5 blastocysts from control ( R26R-mitoAtDarT , upper panel) and R26-mitoAtDarT (lower panel) embryos. Immunofluorescence staining shows SOX2 (inner cell mass marker, green), CDX2 (trophectoderm marker, magenta), and nuclei (DAPI, gray). Images were acquired by light-sheet microscopy and analyzed with Imaris software; the Spots module was used for cell quantification. Scale bars, 20 μm. N = 15 embryos per genotype. B. Quantification of total cell number (DAPI+) and lineage-specific cells (SOX2+ and CDX2+) per blastocyst. R26-mitoAtDarT embryos show significantly fewer total cells and reduced numbers of SOX2+ and CDX2+ cells. Notably, the proportion of cells co-expressing SOX2 and CDX2 is increased in R26-mitoAtDarT embryos. C. A stacked bar chart represents the average of the percentage of CDX2+, SOX2+ and CDX2 and SOX2 double positive cells in R26R-mitoAtDarT and R26-mitoAtDarT. The proportion of double-positive cells, relative to the total cell number, was significantly higher for the different markers, P < 0.002.

Article Snippet: Cryosections of post-implantation embryos were washed with 0.3% PBST three times for 20 minutes each to ensure permeabilization and complete removal of the OCT. Tissues were then blocked in 10% heat-inactivated goat serum (Himedia) and 1% BSA (Sigma) in 0.3 % PBST for 1 hour, followed by overnight incubation with primary antibodies: CdX2 (Biogenix-MU392A), SOX2 (Invitrogen-14-9811-82), SOX17 (R&D systems-af1924), Brachyury (Abcam-EPR18113), TOMM20 (Thermofisher-MA532148), FLAG-tag (Sigma-F1804), phospho-Histone H3 (pHH3) (Cell Signalling Technology - 9706S), and Cleaved caspase (Cell Signalling Technology - 9661S) at a 1:200 concentration diluted in the blocking buffer, overnight at 4°C.

Techniques: Control, Immunofluorescence, Staining, Marker, Microscopy, Software, Expressing

Journal: Journal of Cellular and Molecular Medicine

Article Title: CXCL16 and oxLDL are induced in the onset of diabetic nephropathy

doi: 10.1111/j.1582-4934.2009.00761.x

Figure Lengend Snippet: Effect of HG on cellular CXCL16 and ADAM10 expression in human podocytes. (A) Immunofluorescence staining of CXCL16 and ADAM10 expression in human podocytes treated for 24 or 120 hrs with LG (5 mM) or HG (30 mM) containing media. The cells were stained with DAPI to visualize nuclei (blue), and with Alexa Fluor 488 and Cy3 coupled secondary antibodies to visualize the distribution of ADAM10 (green) and CXCL16 (red) proteins. (B) Immunofluorescence results of CXCL16 and ADAM10 expression after 24 hrs glucose treatment using the BZ-Analyzer software (Keyence). Data represent mean ± S.D. *** P < 0.001, ** P < 0.01 considered statistically significant compared to cells treated with LG concentration. (C) Podocytes were incubated 72 hrs with 5 or 30 mM glucose containing media. Cells were than lysed and ADAM10 expression was determined by Western blot analysis. β-actin was used to demonstrate equal sample loading. (D) CXCL16 protein expression was measured with a CXCL16 specific ELISA 24 hrs after the treatment of podocytes with 5 mM glucose or 30 mM glucose. Data represent mean ± S.D. *** P < 0.001, ** P < 0.01 considered statistically significant compared to cells treated with LG containing media. (E) mRNA from podocytes was isolated at times indicated, transcribed in the cDNA as described in material and methods and real-time PCR was performed. (F) Release of CXCL16 was measured in podocytes incubated for 24, 72 or 120 hrs with LG (5 mM) or HG (30 mM) containing media. Supernatants were prepared as described in material and methods and soluble CXCL16 was measured by CXCL16 specific ELISA. (G-I) ADAM10 expression was down-regulated in human podocytes by ADAM10 specific siRNA (controlled by ADAM10 western blot, depicted above the graph) and cellular (G) and soluble CXCL16 (I) was measured by a CXCL16 specific ELISA. A10 = ADAM10, sc = scrambled.

Article Snippet: The blocking anti-human CXCL16 antibody, recombinant human CXCL16, recombinant human TGF-β and the ADAM10 antibody used for immunofluorescence staining were obtained from R&D Systems (Wiesbaden, Germany).

Techniques: Expressing, Immunofluorescence, Staining, Software, Concentration Assay, Incubation, Western Blot, Enzyme-linked Immunosorbent Assay, Isolation, Real-time Polymerase Chain Reaction

Journal: Journal of Cellular and Molecular Medicine

Article Title: CXCL16 and oxLDL are induced in the onset of diabetic nephropathy

doi: 10.1111/j.1582-4934.2009.00761.x

Figure Lengend Snippet: Expression of CXCL16 and ADAM10 in tissue section of normal kidney and patients with DN. (A) Renal tissue of normal kidney (normal) and of patients diagnosed with DN were analysed with double immunofluorescence staining of CXCL16 (red fluorescence) and ADAM10 (green fluorescence) in glomeruli (upper panel) and tubules (lower panel). (B) Semi-quantitative analysis of fluorescence intensities of glomerular CXCL16 (grey bars) and glomerular and ADAM10 (black bars) expression is depicted in a graph. (C) oxLDL expression in renal sections of normal kidney (normal) and patients diagnosed with DN. Tissue sections were stained with oxLDL antibody followed by Cy3 coupled secondary antibody (red). (D) Immunofluorescence results of oxLDL expression in renal tissue of normal kidney and of patients with DN. Data represent mean ± S.D. *** P < 0.001, ** P < 0.01 considered statistically significant compared to the normal kidney. (E) Double immunofluorescense analysis of oxLDL (red colour) and CXCL16 (green colour) was performed on kidney biopsies of patients with DN. Tissue section was stained with DAPI to visualize the nuclei of the cells.

Article Snippet: The blocking anti-human CXCL16 antibody, recombinant human CXCL16, recombinant human TGF-β and the ADAM10 antibody used for immunofluorescence staining were obtained from R&D Systems (Wiesbaden, Germany).

Techniques: Expressing, Double Immunofluorescence Staining, Fluorescence, Staining, Immunofluorescence

Journal: International Journal of Oncology

Article Title: Expression and clinical significance of CXCR5/CXCL13 in human non-small cell lung carcinoma

doi: 10.3892/ijo.2014.2688

Figure Lengend Snippet: Immunohistochemical detection of CXCR5 in lung cancer (LuCa) tissues. Representative figures of (A) non-neoplastic (n=8), (B) squamous cell carcinoma (SCC) (n=24) and (C) adenocarcinoma (AC) (n=54) lung tissues stained with anti-CXCR5 antibodies. Brown [3,3′-diaminobenzidine (DAB)] color shows CXCR5 staining. The images were captured using an Aperio ScanScope CS system with a 40× objective. Immuno-intensities of CXCR5 in each section were quantified using image analysis Aperio ImageScope v.6.25 software.

Article Snippet: Next, NCI-H1915 and SW-1271 cells were incubated with isotype control or anti-human CXCR5 antibody at a concentration of 1 μg/ml (both from R&D Systems) for 1 h at 37°C with 5% CO 2 and added to the top chambers in serum-free RPMI medium at 10,000 cells per well.

Techniques: Immunohistochemical staining, Staining, Software

Journal: International Journal of Oncology

Article Title: Expression and clinical significance of CXCR5/CXCL13 in human non-small cell lung carcinoma

doi: 10.3892/ijo.2014.2688

Figure Lengend Snippet: CXCR5 expression by lung cancer (LuCa) tissues. (A) Representative figures of non-neoplastic (n=8), squamous cell carcinoma (SCC) (n=22), and adenocarcinoma (AC) (n=52) lung tissues stained with isotype control or anti-CXCR5 antibodies. Positive pixel counts were quantified using a membrane algorithm of image analysis Aperio ImageScope v.6.25 software. (B) The average positive pixel count of CXCR5, in non-neoplastic (n=8), SCC (n=22), and AC (n=52) tissues, quantified with a nuclear algorithm of image analysis Aperio ImageScope v.6.25 software. * Significant differences (p<0.001) between groups with LuCa and control. (C–F) Average pixel counts for CXCR5 for different tumor stages and for tumors with nodal involvement in SCC and AC cases, respectively.

Article Snippet: Next, NCI-H1915 and SW-1271 cells were incubated with isotype control or anti-human CXCR5 antibody at a concentration of 1 μg/ml (both from R&D Systems) for 1 h at 37°C with 5% CO 2 and added to the top chambers in serum-free RPMI medium at 10,000 cells per well.

Techniques: Expressing, Staining, Control, Membrane, Software

Journal: International Journal of Oncology

Article Title: Expression and clinical significance of CXCR5/CXCL13 in human non-small cell lung carcinoma

doi: 10.3892/ijo.2014.2688

Figure Lengend Snippet: Membrane CXCR5 expression by lung cancer (LuCa) tissues. (A) Representative figure of non-neoplastic (n=8), squamous cell carcinoma (SCC) (n=22) and adenocarcinoma (AC) (n=52) lung tissues stained with isotype control or anti-CXCR5 antibodies. Brown [3,3′-diaminobenzidine (DAB)] color show CXCR5 staining. An Aperio ScanScope CS system with a 40× objective captured digital images of each slide. The membrane intensity of CXCR5 was quantified using a membrane algorithm of image analysis Aperio ImageScope v.6.25 software. (B) The membrane intensity of CXCR5, in non-neoplastic (n=8), SCC (n=22), and AC (n=52) tissues, quantified using a nuclear algorithm of image analysis Aperio ImageScope v.6.25 software. * Significant differences (p<0.001) between groups with LuCa and control. (C–F) Membrane intensities of CXCR5 for different tumor stages and for tumors with nodal involvement in SCC and AC cases, respectively.

Article Snippet: Next, NCI-H1915 and SW-1271 cells were incubated with isotype control or anti-human CXCR5 antibody at a concentration of 1 μg/ml (both from R&D Systems) for 1 h at 37°C with 5% CO 2 and added to the top chambers in serum-free RPMI medium at 10,000 cells per well.

Techniques: Membrane, Expressing, Staining, Control, Software

Journal: International Journal of Oncology

Article Title: Expression and clinical significance of CXCR5/CXCL13 in human non-small cell lung carcinoma

doi: 10.3892/ijo.2014.2688

Figure Lengend Snippet: Nuclear CXCR5 expression in lung cancer (LuCa) tissues. (A) Representative figures of non-neoplastic (n=8), squamous cell carcinoma (SCC) (n=22), and adenocarcinoma (AC) (n=52) lung tissues stained with isotype control or anti-CXCR5 antibodies. Brown [3,3′-diaminobenzidine (DAB)] color show CXCR5 staining. An Aperio ScanScope CS system with a 40× objective captured digital images of each slide. Stained cells with negative and positive nuclei were counted and categorized according to stain intensity 0 (Blue), 1+(yellow), 2+(orange) and 3+(Red). (B) The nuclear intensity of CXCR5, in non-neoplastic (n=8), SCC (n=22), and AC (n=52) tissues, quantified using a nuclear algorithm of image analysis Aperio ImageScope v.6.25 software. * Significant differences (p<0.001) between groups with LuCa and control. (C–F) Nuclear intensities of CXCR5 in different tumor stages and in tumors with nodal involvement in SCC and AC cases, respectively.

Article Snippet: Next, NCI-H1915 and SW-1271 cells were incubated with isotype control or anti-human CXCR5 antibody at a concentration of 1 μg/ml (both from R&D Systems) for 1 h at 37°C with 5% CO 2 and added to the top chambers in serum-free RPMI medium at 10,000 cells per well.

Techniques: Expressing, Staining, Control, Software

Journal: International Journal of Oncology

Article Title: Expression and clinical significance of CXCR5/CXCL13 in human non-small cell lung carcinoma

doi: 10.3892/ijo.2014.2688

Figure Lengend Snippet: Expression of CXCR5 in lung cancer (LuCa) cells. (A) Non-small cell lung carcinoma (NCI-H1915) and small cell lung carcinoma (SW-1271) cells were stained with FITC-conjugated isotype control antibodies (solid histogram) or FITC-conjugated anti-CXCR5 (open histogram) and quantified in triplicates by flow cytometry. The experiments were repeated three times. (B) NCI-H1915 and SW-1271 cells were stained with FITC-conjugated anti-CXCR5 or FITC-conjugated isotype control antibodies. Images were acquired by multispectral imaging flow cytometry.

Article Snippet: Next, NCI-H1915 and SW-1271 cells were incubated with isotype control or anti-human CXCR5 antibody at a concentration of 1 μg/ml (both from R&D Systems) for 1 h at 37°C with 5% CO 2 and added to the top chambers in serum-free RPMI medium at 10,000 cells per well.

Techniques: Expressing, Staining, Control, Flow Cytometry, Imaging

Journal: International Journal of Oncology

Article Title: Expression and clinical significance of CXCR5/CXCL13 in human non-small cell lung carcinoma

doi: 10.3892/ijo.2014.2688

Figure Lengend Snippet: CXCL13 mediated lung cancer (LuCa) cell migration. NSCLC (NCI-H1915) and SCLC (SW-1271) cells were tested for their capacity to migrate toward the chemotactic gradients of 0 (open boxes) or 100 ng/ml of CXCL13 (closed boxes). One group of cells was treated with anti-human CXCR5 antibody (1 μg/ml, hashed boxes) before the migration assay. * Significant differences (p<0.01) between no addition and addition and chemokine-induced cells. # Significant difference between chemokine engagement and receptor neutralization.

Article Snippet: Next, NCI-H1915 and SW-1271 cells were incubated with isotype control or anti-human CXCR5 antibody at a concentration of 1 μg/ml (both from R&D Systems) for 1 h at 37°C with 5% CO 2 and added to the top chambers in serum-free RPMI medium at 10,000 cells per well.

Techniques: Migration, Neutralization