heat-inactivated fbs Search Results


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  • 99
    Millipore heat inactivated fbs
    F4/80 + CD11b + Gr1 + (Subset 1) cells accumulated at the mucosal damaged area in early phase of healing. 1 × 10 7 BMC were labeled using CFDA-SE. These cells were injected into AA + 5-FU mice via tail vein. After 24 hours, colon was dissected and incubated in calcium and magnesium-free <t>HBSS</t> containing 2.5% heat-inactivated <t>FBS</t> and 1 mM DTT to remove mucus. After the tissues were treated with collagenase and DNase I for 60 min at 37°C, cells were washed and stained with 7-AAD, F4/80-APC, and C11b-PE or 7-AAD, F4/80-APC, and Gr1-PE, for the flow cytometric analysis. 7-AAD was used for exclusion of dead cells. (a) CFDA-SE positive cells indicated bone marrow-derived cells. (b) CFDA-SE positive population was CD11b + F4/80 + monocytes. (c) F4/80 positive population was almost all Gr1 + cells, indicating the F4/80 + Gr1 + Subset 1 monocytes. (d) The population had typical monocytes morphology.
    Heat Inactivated Fbs, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 2938 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    97
    GE Healthcare heat inactivated fbs
    Protein coronas modulate sulphide formation. Proposed mechanism of protein corona-modulated nano-Ag 2 S formation at Ag <t>NPs,</t> with hard corona proteins trapping Ag + released from the nanoparticle surface and soft corona proteins transporting said ions away from the sulphide-formation centres in the long-lived corona ( a ); TEM images of silver nanocubes after 24 h in RPMI-1640 cell culture medium supplemented with 1% <t>FBS</t> ( b ), followed by 6 days incubation in RPMI-1640 with 0% FBS (inset cartoon showing only hard corona around Ag NPs) ( c ), 1% FBS ( d ) or 10% FBS ( e ; common inset cartoon showing hard and soft coronas, as well as free bulk proteins around Ag NPs); TEM images of silver nanocubes after 7 days incubation in RPMI-1640 with 0.4 mg ml −1 BSA ( f ) or 4 mg ml −1 BSA ( g ) (common inset cartoon showing hard corona and free bulk proteins around Ag NPs); Ultraviolet–visible spectra of cubic ( h ) and quasi-spherical ( i ) Ag NPs after 24 h incubation in RPMI-1640 cell culture medium supplemented with 1, 10 or 50% FBS; TEM images of silver nanocubes after 24 h in RPMI-1640 supplemented with 1% FBS ( j ), 10% FBS ( k ) and 50% FBS ( l ). Scale bars are 100 nm ( b , j – l ) or 50 nm ( c – g ).
    Heat Inactivated Fbs, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 97/100, based on 2186 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Atlanta Biologicals heat inactivated fbs
    Inflammatory gene expression profile in PGRN pretreated microglia. BV2 microglia were plated at 150,000 cells per well in a 12-well plate in <t>DMEM/F-12</t> with 5% <t>FBS.</t> Cells were treated for 16 h with saline followed by 24 h of saline (Control), or saline
    Heat Inactivated Fbs, supplied by Atlanta Biologicals, used in various techniques. Bioz Stars score: 94/100, based on 557 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Biochrom heat inactivated fbs
    Inflammatory gene expression profile in PGRN pretreated microglia. BV2 microglia were plated at 150,000 cells per well in a 12-well plate in <t>DMEM/F-12</t> with 5% <t>FBS.</t> Cells were treated for 16 h with saline followed by 24 h of saline (Control), or saline
    Heat Inactivated Fbs, supplied by Biochrom, used in various techniques. Bioz Stars score: 92/100, based on 143 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Cellgro heat inactivated fbs
    Activation of CD271 by nerve growth factor confers a more invasive phenotype in MOC2 cells and HSC3 cells by the induction of Snai2/Slug expression. (A-B) Comparison of cancer cell invasiveness in response to recombinant human nerve growth factor-β (rhNGF-β) treatment. Growth factor reduced Matrigel® Invasion Chambers were used to assess and compare the invasive potential of MOC2 vs. MOC2-CD271 cells (A), and that of HSC3 vs. HSC3-CD271 cells (B). Cells were cultured in low serum media <t>(DMEM/F12</t> with 2% fetal bovine serum) for 24 hrs, and then, treated with rhNGF-β (100ng/mL). Cells were collected and incubated in invasion chambers with fibronectin (10ng/mL) as a chemoattractant for 48 hrs. Quantitation of invasion assay was performed by counting invaded cells in 6 random fields per chamber under light microscopy. Mean±SD is shown, **p
    Heat Inactivated Fbs, supplied by Cellgro, used in various techniques. Bioz Stars score: 92/100, based on 175 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    GenDEPOT heat inactivated fbs
    Activation of CD271 by nerve growth factor confers a more invasive phenotype in MOC2 cells and HSC3 cells by the induction of Snai2/Slug expression. (A-B) Comparison of cancer cell invasiveness in response to recombinant human nerve growth factor-β (rhNGF-β) treatment. Growth factor reduced Matrigel® Invasion Chambers were used to assess and compare the invasive potential of MOC2 vs. MOC2-CD271 cells (A), and that of HSC3 vs. HSC3-CD271 cells (B). Cells were cultured in low serum media <t>(DMEM/F12</t> with 2% fetal bovine serum) for 24 hrs, and then, treated with rhNGF-β (100ng/mL). Cells were collected and incubated in invasion chambers with fibronectin (10ng/mL) as a chemoattractant for 48 hrs. Quantitation of invasion assay was performed by counting invaded cells in 6 random fields per chamber under light microscopy. Mean±SD is shown, **p
    Heat Inactivated Fbs, supplied by GenDEPOT, used in various techniques. Bioz Stars score: 90/100, based on 12 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Wisent Corporation heat inactivated fbs
    Activation of CD271 by nerve growth factor confers a more invasive phenotype in MOC2 cells and HSC3 cells by the induction of Snai2/Slug expression. (A-B) Comparison of cancer cell invasiveness in response to recombinant human nerve growth factor-β (rhNGF-β) treatment. Growth factor reduced Matrigel® Invasion Chambers were used to assess and compare the invasive potential of MOC2 vs. MOC2-CD271 cells (A), and that of HSC3 vs. HSC3-CD271 cells (B). Cells were cultured in low serum media <t>(DMEM/F12</t> with 2% fetal bovine serum) for 24 hrs, and then, treated with rhNGF-β (100ng/mL). Cells were collected and incubated in invasion chambers with fibronectin (10ng/mL) as a chemoattractant for 48 hrs. Quantitation of invasion assay was performed by counting invaded cells in 6 random fields per chamber under light microscopy. Mean±SD is shown, **p
    Heat Inactivated Fbs, supplied by Wisent Corporation, used in various techniques. Bioz Stars score: 94/100, based on 141 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    Cytogen heat inactivated fbs
    Activation of CD271 by nerve growth factor confers a more invasive phenotype in MOC2 cells and HSC3 cells by the induction of Snai2/Slug expression. (A-B) Comparison of cancer cell invasiveness in response to recombinant human nerve growth factor-β (rhNGF-β) treatment. Growth factor reduced Matrigel® Invasion Chambers were used to assess and compare the invasive potential of MOC2 vs. MOC2-CD271 cells (A), and that of HSC3 vs. HSC3-CD271 cells (B). Cells were cultured in low serum media <t>(DMEM/F12</t> with 2% fetal bovine serum) for 24 hrs, and then, treated with rhNGF-β (100ng/mL). Cells were collected and incubated in invasion chambers with fibronectin (10ng/mL) as a chemoattractant for 48 hrs. Quantitation of invasion assay was performed by counting invaded cells in 6 random fields per chamber under light microscopy. Mean±SD is shown, **p
    Heat Inactivated Fbs, supplied by Cytogen, used in various techniques. Bioz Stars score: 90/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Equitech-Bio heat inactivated fbs
    Loss of GADD34 increased cellular damage and apoptosis in <t>RAW</t> 264.7 cells treated with LPS and Tyr/Cys-deprivation. (a). Knockdown of GADD34 by shRNA was evaluated. Real-time PCR analysis of GADD34 expression in GADD34-deficient (shGADD34) and control RAW 264.7 cells (shControl) after treatment with LPS and Tyr/Cys-deprivation for 8 h. Graph shows the relative expression as means ± SE of three independent experiments. (b). GADD34-deficient or control RAW 264.7 cells were treated with DMEM + <t>10%FBS</t> (control) or LPS (1 μg/mL) in Tyr/Cys-deprivation medium for 8 h or 14 h. Cells were stained with 7-AAD and PE-labeled Annexin V and assessed by flow cytometry. (c). GADD34-deficient or control RAW 264.7 cells were treated with LPS with or without Tyr/Cys-deprivation for 14 h. Cells were stained with 7-AAD and PE-labeled Annexin V and assessed by flow cytometry. Relative % of Annexin V + 7AAD − apoptotic cells shown as means ± SE of three independent experiments. (d). BMDMs from GADD34 KO or WT were treated with LPS (1 μg/mL) combined with Tyr/Cys-deprivation for 8 h. Cells were stained with 7-AAD and PE-labeled Annexin V and assessed by flow cytometry. (e). GADD34-deficient or control RAW 264.7 cells were treated with LPS (+L; 1 μg/mL) and/or Tyr/Cys-deprivation for 24 h. Cell lysates were immunoblotted with anti-caspase 3 antibody and detected full length caspase 3 (35 kDa) and cleaved caspase 3 (17, 19 kDa). (f). GADD34-deficient or control RAW 264.7 cells were treated with DMEM + 10% FBS (control) or LPS (1 μg/mL) with or without Tyr/Cys-deprivation medium for 24 h. After treatment, cells were collected and counted by trypan-blue dye exclusion using a Burker-Turk cell count chamber. Data are means ± SE of cell number from three independent experiments. Data are representative of three independent experiments (b, d, e). * p
    Heat Inactivated Fbs, supplied by Equitech-Bio, used in various techniques. Bioz Stars score: 93/100, based on 52 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Lonza heat inactivated fbs
    Loss of GADD34 increased cellular damage and apoptosis in <t>RAW</t> 264.7 cells treated with LPS and Tyr/Cys-deprivation. (a). Knockdown of GADD34 by shRNA was evaluated. Real-time PCR analysis of GADD34 expression in GADD34-deficient (shGADD34) and control RAW 264.7 cells (shControl) after treatment with LPS and Tyr/Cys-deprivation for 8 h. Graph shows the relative expression as means ± SE of three independent experiments. (b). GADD34-deficient or control RAW 264.7 cells were treated with DMEM + <t>10%FBS</t> (control) or LPS (1 μg/mL) in Tyr/Cys-deprivation medium for 8 h or 14 h. Cells were stained with 7-AAD and PE-labeled Annexin V and assessed by flow cytometry. (c). GADD34-deficient or control RAW 264.7 cells were treated with LPS with or without Tyr/Cys-deprivation for 14 h. Cells were stained with 7-AAD and PE-labeled Annexin V and assessed by flow cytometry. Relative % of Annexin V + 7AAD − apoptotic cells shown as means ± SE of three independent experiments. (d). BMDMs from GADD34 KO or WT were treated with LPS (1 μg/mL) combined with Tyr/Cys-deprivation for 8 h. Cells were stained with 7-AAD and PE-labeled Annexin V and assessed by flow cytometry. (e). GADD34-deficient or control RAW 264.7 cells were treated with LPS (+L; 1 μg/mL) and/or Tyr/Cys-deprivation for 24 h. Cell lysates were immunoblotted with anti-caspase 3 antibody and detected full length caspase 3 (35 kDa) and cleaved caspase 3 (17, 19 kDa). (f). GADD34-deficient or control RAW 264.7 cells were treated with DMEM + 10% FBS (control) or LPS (1 μg/mL) with or without Tyr/Cys-deprivation medium for 24 h. After treatment, cells were collected and counted by trypan-blue dye exclusion using a Burker-Turk cell count chamber. Data are means ± SE of cell number from three independent experiments. Data are representative of three independent experiments (b, d, e). * p
    Heat Inactivated Fbs, supplied by Lonza, used in various techniques. Bioz Stars score: 93/100, based on 275 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Cambrex heat inactivated fbs
    Loss of GADD34 increased cellular damage and apoptosis in <t>RAW</t> 264.7 cells treated with LPS and Tyr/Cys-deprivation. (a). Knockdown of GADD34 by shRNA was evaluated. Real-time PCR analysis of GADD34 expression in GADD34-deficient (shGADD34) and control RAW 264.7 cells (shControl) after treatment with LPS and Tyr/Cys-deprivation for 8 h. Graph shows the relative expression as means ± SE of three independent experiments. (b). GADD34-deficient or control RAW 264.7 cells were treated with DMEM + <t>10%FBS</t> (control) or LPS (1 μg/mL) in Tyr/Cys-deprivation medium for 8 h or 14 h. Cells were stained with 7-AAD and PE-labeled Annexin V and assessed by flow cytometry. (c). GADD34-deficient or control RAW 264.7 cells were treated with LPS with or without Tyr/Cys-deprivation for 14 h. Cells were stained with 7-AAD and PE-labeled Annexin V and assessed by flow cytometry. Relative % of Annexin V + 7AAD − apoptotic cells shown as means ± SE of three independent experiments. (d). BMDMs from GADD34 KO or WT were treated with LPS (1 μg/mL) combined with Tyr/Cys-deprivation for 8 h. Cells were stained with 7-AAD and PE-labeled Annexin V and assessed by flow cytometry. (e). GADD34-deficient or control RAW 264.7 cells were treated with LPS (+L; 1 μg/mL) and/or Tyr/Cys-deprivation for 24 h. Cell lysates were immunoblotted with anti-caspase 3 antibody and detected full length caspase 3 (35 kDa) and cleaved caspase 3 (17, 19 kDa). (f). GADD34-deficient or control RAW 264.7 cells were treated with DMEM + 10% FBS (control) or LPS (1 μg/mL) with or without Tyr/Cys-deprivation medium for 24 h. After treatment, cells were collected and counted by trypan-blue dye exclusion using a Burker-Turk cell count chamber. Data are means ± SE of cell number from three independent experiments. Data are representative of three independent experiments (b, d, e). * p
    Heat Inactivated Fbs, supplied by Cambrex, used in various techniques. Bioz Stars score: 92/100, based on 24 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Gemini Bio heat inactivated fbs
    QD232 inhibits cell migration. (A) A 24 h treatment with QD232 (1 μM) resulted in decreased migration of serum-starved <t>MIA</t> PaCa-2 cells through the membrane of a Boyden chamber in the presence of 10% <t>FBS</t> stimulation. Cells were imaged after fixing with methanol and staining with Giemsa using a Nikon microscope with 10× objective. (B) Quantification of data (means ± SD) shown in (A); effect induced by both concentrations of QD232 were significantly different from control (10% FBS only); P ). All concentrations of QD232, except the lowest (0.05μM in F, and 0.05 and 0.1 μM in G), had significant effects, compared with 10% FBS only; P
    Heat Inactivated Fbs, supplied by Gemini Bio, used in various techniques. Bioz Stars score: 93/100, based on 209 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Omega Scientific Inc heat inactivated fbs
    QD232 inhibits cell migration. (A) A 24 h treatment with QD232 (1 μM) resulted in decreased migration of serum-starved <t>MIA</t> PaCa-2 cells through the membrane of a Boyden chamber in the presence of 10% <t>FBS</t> stimulation. Cells were imaged after fixing with methanol and staining with Giemsa using a Nikon microscope with 10× objective. (B) Quantification of data (means ± SD) shown in (A); effect induced by both concentrations of QD232 were significantly different from control (10% FBS only); P ). All concentrations of QD232, except the lowest (0.05μM in F, and 0.05 and 0.1 μM in G), had significant effects, compared with 10% FBS only; P
    Heat Inactivated Fbs, supplied by Omega Scientific Inc, used in various techniques. Bioz Stars score: 93/100, based on 164 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    PAA Laboratories heat inactivated fbs
    QD232 inhibits cell migration. (A) A 24 h treatment with QD232 (1 μM) resulted in decreased migration of serum-starved <t>MIA</t> PaCa-2 cells through the membrane of a Boyden chamber in the presence of 10% <t>FBS</t> stimulation. Cells were imaged after fixing with methanol and staining with Giemsa using a Nikon microscope with 10× objective. (B) Quantification of data (means ± SD) shown in (A); effect induced by both concentrations of QD232 were significantly different from control (10% FBS only); P ). All concentrations of QD232, except the lowest (0.05μM in F, and 0.05 and 0.1 μM in G), had significant effects, compared with 10% FBS only; P
    Heat Inactivated Fbs, supplied by PAA Laboratories, used in various techniques. Bioz Stars score: 92/100, based on 57 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Biological Industries Inc heat inactivated fbs
    QD232 inhibits cell migration. (A) A 24 h treatment with QD232 (1 μM) resulted in decreased migration of serum-starved <t>MIA</t> PaCa-2 cells through the membrane of a Boyden chamber in the presence of 10% <t>FBS</t> stimulation. Cells were imaged after fixing with methanol and staining with Giemsa using a Nikon microscope with 10× objective. (B) Quantification of data (means ± SD) shown in (A); effect induced by both concentrations of QD232 were significantly different from control (10% FBS only); P ). All concentrations of QD232, except the lowest (0.05μM in F, and 0.05 and 0.1 μM in G), had significant effects, compared with 10% FBS only; P
    Heat Inactivated Fbs, supplied by Biological Industries Inc, used in various techniques. Bioz Stars score: 94/100, based on 117 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Biowest SAS heat inactivated fbs
    QD232 inhibits cell migration. (A) A 24 h treatment with QD232 (1 μM) resulted in decreased migration of serum-starved <t>MIA</t> PaCa-2 cells through the membrane of a Boyden chamber in the presence of 10% <t>FBS</t> stimulation. Cells were imaged after fixing with methanol and staining with Giemsa using a Nikon microscope with 10× objective. (B) Quantification of data (means ± SD) shown in (A); effect induced by both concentrations of QD232 were significantly different from control (10% FBS only); P ). All concentrations of QD232, except the lowest (0.05μM in F, and 0.05 and 0.1 μM in G), had significant effects, compared with 10% FBS only; P
    Heat Inactivated Fbs, supplied by Biowest SAS, used in various techniques. Bioz Stars score: 94/100, based on 145 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    EuroClone heat inactivated fbs
    Effect of ST258 KP strains on inflammatory cytokines production by monocytes and MDC. Monocytes or MDC were cultured at 10 6 cells/ml in <t>RPMI</t> with 10% <t>FBS</t> in the presence or absence (US) of UV-inactivated ST258-KP strains at 1:10 ratio or of LPS (400 ng/ml). Conditioned media were collected after 16 hours of culture. Cytokine production was measured by Immunoplex array. Box-chart plots show cytokine production from 6 different healthy donors. The boxes extend from SE; the horizontal line represents the median, asterisks indicate the mean value. Statistical analysis was performed by Student’s t-test and p ≤ 0.05 was considered significant.
    Heat Inactivated Fbs, supplied by EuroClone, used in various techniques. Bioz Stars score: 92/100, based on 130 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Welgene inc heat inactivated fbs
    Effect of ST258 KP strains on inflammatory cytokines production by monocytes and MDC. Monocytes or MDC were cultured at 10 6 cells/ml in <t>RPMI</t> with 10% <t>FBS</t> in the presence or absence (US) of UV-inactivated ST258-KP strains at 1:10 ratio or of LPS (400 ng/ml). Conditioned media were collected after 16 hours of culture. Cytokine production was measured by Immunoplex array. Box-chart plots show cytokine production from 6 different healthy donors. The boxes extend from SE; the horizontal line represents the median, asterisks indicate the mean value. Statistical analysis was performed by Student’s t-test and p ≤ 0.05 was considered significant.
    Heat Inactivated Fbs, supplied by Welgene inc, used in various techniques. Bioz Stars score: 94/100, based on 73 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Flow Laboratories heat inactivated fbs
    Effect of ST258 KP strains on inflammatory cytokines production by monocytes and MDC. Monocytes or MDC were cultured at 10 6 cells/ml in <t>RPMI</t> with 10% <t>FBS</t> in the presence or absence (US) of UV-inactivated ST258-KP strains at 1:10 ratio or of LPS (400 ng/ml). Conditioned media were collected after 16 hours of culture. Cytokine production was measured by Immunoplex array. Box-chart plots show cytokine production from 6 different healthy donors. The boxes extend from SE; the horizontal line represents the median, asterisks indicate the mean value. Statistical analysis was performed by Student’s t-test and p ≤ 0.05 was considered significant.
    Heat Inactivated Fbs, supplied by Flow Laboratories, used in various techniques. Bioz Stars score: 92/100, based on 12 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Carlo Erba Reagents heat inactivated fbs
    Effect of ST258 KP strains on inflammatory cytokines production by monocytes and MDC. Monocytes or MDC were cultured at 10 6 cells/ml in <t>RPMI</t> with 10% <t>FBS</t> in the presence or absence (US) of UV-inactivated ST258-KP strains at 1:10 ratio or of LPS (400 ng/ml). Conditioned media were collected after 16 hours of culture. Cytokine production was measured by Immunoplex array. Box-chart plots show cytokine production from 6 different healthy donors. The boxes extend from SE; the horizontal line represents the median, asterisks indicate the mean value. Statistical analysis was performed by Student’s t-test and p ≤ 0.05 was considered significant.
    Heat Inactivated Fbs, supplied by Carlo Erba Reagents, used in various techniques. Bioz Stars score: 93/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    LabTech Inc heat inactivated fbs
    Effect of ST258 KP strains on inflammatory cytokines production by monocytes and MDC. Monocytes or MDC were cultured at 10 6 cells/ml in <t>RPMI</t> with 10% <t>FBS</t> in the presence or absence (US) of UV-inactivated ST258-KP strains at 1:10 ratio or of LPS (400 ng/ml). Conditioned media were collected after 16 hours of culture. Cytokine production was measured by Immunoplex array. Box-chart plots show cytokine production from 6 different healthy donors. The boxes extend from SE; the horizontal line represents the median, asterisks indicate the mean value. Statistical analysis was performed by Student’s t-test and p ≤ 0.05 was considered significant.
    Heat Inactivated Fbs, supplied by LabTech Inc, used in various techniques. Bioz Stars score: 90/100, based on 57 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Biowhittaker Inc heat inactivated fbs
    Effect of ST258 KP strains on inflammatory cytokines production by monocytes and MDC. Monocytes or MDC were cultured at 10 6 cells/ml in <t>RPMI</t> with 10% <t>FBS</t> in the presence or absence (US) of UV-inactivated ST258-KP strains at 1:10 ratio or of LPS (400 ng/ml). Conditioned media were collected after 16 hours of culture. Cytokine production was measured by Immunoplex array. Box-chart plots show cytokine production from 6 different healthy donors. The boxes extend from SE; the horizontal line represents the median, asterisks indicate the mean value. Statistical analysis was performed by Student’s t-test and p ≤ 0.05 was considered significant.
    Heat Inactivated Fbs, supplied by Biowhittaker Inc, used in various techniques. Bioz Stars score: 92/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    CPX does not alter mRNA expression, but inhibits protein synthesis and promotes protein degradation of VEGFR-3 A, CPX did not affect VEGFR-3 mRNA level. Total RNA was extracted from LECs treated with CPX (0-5 μM) for 24 h (Left panel) or with CPX (5 μM) for 0-24 h (Right panel), followed by semi-quantitative RT-PCR. β-actin was used as a loading control. B, CPX inhibited protein synthesis of VEGFR-3 in LECs. LECs were pretreated with CPX (0-5 μM) for 24 h (Left panel) or with CPX (5 μM) for 0-24 h (Right panel), and then pulsed with 35 S-Met/Cys for 4 h, followed by immunoprecipitation with antibodies to VEGFR-3. The immunoprecipitates were separated by SDS-PAGE and transferred to PVDF membranes, followed by autoradiography. GAPDH served as an internal control. C, CPX promoted protein degradation of VEGFR-3 in LECs. LECs, grown in 10% <t>FBS-DMEM</t> medium, were exposed to cycloheximide (CHX, 50 μg/ml), in the presence or absence of CPX (5 μM), for 0-12 h, followed by Western blot analysis with indicated antibodies.
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    CPX does not alter mRNA expression, but inhibits protein synthesis and promotes protein degradation of VEGFR-3 A, CPX did not affect VEGFR-3 mRNA level. Total RNA was extracted from LECs treated with CPX (0-5 μM) for 24 h (Left panel) or with CPX (5 μM) for 0-24 h (Right panel), followed by semi-quantitative RT-PCR. β-actin was used as a loading control. B, CPX inhibited protein synthesis of VEGFR-3 in LECs. LECs were pretreated with CPX (0-5 μM) for 24 h (Left panel) or with CPX (5 μM) for 0-24 h (Right panel), and then pulsed with 35 S-Met/Cys for 4 h, followed by immunoprecipitation with antibodies to VEGFR-3. The immunoprecipitates were separated by SDS-PAGE and transferred to PVDF membranes, followed by autoradiography. GAPDH served as an internal control. C, CPX promoted protein degradation of VEGFR-3 in LECs. LECs, grown in 10% <t>FBS-DMEM</t> medium, were exposed to cycloheximide (CHX, 50 μg/ml), in the presence or absence of CPX (5 μM), for 0-12 h, followed by Western blot analysis with indicated antibodies.
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    CPX does not alter mRNA expression, but inhibits protein synthesis and promotes protein degradation of VEGFR-3 A, CPX did not affect VEGFR-3 mRNA level. Total RNA was extracted from LECs treated with CPX (0-5 μM) for 24 h (Left panel) or with CPX (5 μM) for 0-24 h (Right panel), followed by semi-quantitative RT-PCR. β-actin was used as a loading control. B, CPX inhibited protein synthesis of VEGFR-3 in LECs. LECs were pretreated with CPX (0-5 μM) for 24 h (Left panel) or with CPX (5 μM) for 0-24 h (Right panel), and then pulsed with 35 S-Met/Cys for 4 h, followed by immunoprecipitation with antibodies to VEGFR-3. The immunoprecipitates were separated by SDS-PAGE and transferred to PVDF membranes, followed by autoradiography. GAPDH served as an internal control. C, CPX promoted protein degradation of VEGFR-3 in LECs. LECs, grown in 10% <t>FBS-DMEM</t> medium, were exposed to cycloheximide (CHX, 50 μg/ml), in the presence or absence of CPX (5 μM), for 0-12 h, followed by Western blot analysis with indicated antibodies.
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    Image Search Results


    F4/80 + CD11b + Gr1 + (Subset 1) cells accumulated at the mucosal damaged area in early phase of healing. 1 × 10 7 BMC were labeled using CFDA-SE. These cells were injected into AA + 5-FU mice via tail vein. After 24 hours, colon was dissected and incubated in calcium and magnesium-free HBSS containing 2.5% heat-inactivated FBS and 1 mM DTT to remove mucus. After the tissues were treated with collagenase and DNase I for 60 min at 37°C, cells were washed and stained with 7-AAD, F4/80-APC, and C11b-PE or 7-AAD, F4/80-APC, and Gr1-PE, for the flow cytometric analysis. 7-AAD was used for exclusion of dead cells. (a) CFDA-SE positive cells indicated bone marrow-derived cells. (b) CFDA-SE positive population was CD11b + F4/80 + monocytes. (c) F4/80 positive population was almost all Gr1 + cells, indicating the F4/80 + Gr1 + Subset 1 monocytes. (d) The population had typical monocytes morphology.

    Journal: Mediators of Inflammation

    Article Title: Role of Bone Marrow-Derived Monocytes/Macrophages in the Repair of Mucosal Damage Caused by Irradiation and/or Anticancer Drugs in Colitis Model

    doi: 10.1155/2010/634145

    Figure Lengend Snippet: F4/80 + CD11b + Gr1 + (Subset 1) cells accumulated at the mucosal damaged area in early phase of healing. 1 × 10 7 BMC were labeled using CFDA-SE. These cells were injected into AA + 5-FU mice via tail vein. After 24 hours, colon was dissected and incubated in calcium and magnesium-free HBSS containing 2.5% heat-inactivated FBS and 1 mM DTT to remove mucus. After the tissues were treated with collagenase and DNase I for 60 min at 37°C, cells were washed and stained with 7-AAD, F4/80-APC, and C11b-PE or 7-AAD, F4/80-APC, and Gr1-PE, for the flow cytometric analysis. 7-AAD was used for exclusion of dead cells. (a) CFDA-SE positive cells indicated bone marrow-derived cells. (b) CFDA-SE positive population was CD11b + F4/80 + monocytes. (c) F4/80 positive population was almost all Gr1 + cells, indicating the F4/80 + Gr1 + Subset 1 monocytes. (d) The population had typical monocytes morphology.

    Article Snippet: Preparation of Damaged Mucosa and Identification of Injected BMCs Dissected mucosa was incubated in calcium and magnesium-free HBSS (GIBCO) containing 2.5% heat-inactivated FBS and 1 mM DTT (Sigma-Aldrich) to remove mucus.

    Techniques: Labeling, Injection, Mouse Assay, Incubation, Staining, Flow Cytometry, Derivative Assay

    Protein coronas modulate sulphide formation. Proposed mechanism of protein corona-modulated nano-Ag 2 S formation at Ag NPs, with hard corona proteins trapping Ag + released from the nanoparticle surface and soft corona proteins transporting said ions away from the sulphide-formation centres in the long-lived corona ( a ); TEM images of silver nanocubes after 24 h in RPMI-1640 cell culture medium supplemented with 1% FBS ( b ), followed by 6 days incubation in RPMI-1640 with 0% FBS (inset cartoon showing only hard corona around Ag NPs) ( c ), 1% FBS ( d ) or 10% FBS ( e ; common inset cartoon showing hard and soft coronas, as well as free bulk proteins around Ag NPs); TEM images of silver nanocubes after 7 days incubation in RPMI-1640 with 0.4 mg ml −1 BSA ( f ) or 4 mg ml −1 BSA ( g ) (common inset cartoon showing hard corona and free bulk proteins around Ag NPs); Ultraviolet–visible spectra of cubic ( h ) and quasi-spherical ( i ) Ag NPs after 24 h incubation in RPMI-1640 cell culture medium supplemented with 1, 10 or 50% FBS; TEM images of silver nanocubes after 24 h in RPMI-1640 supplemented with 1% FBS ( j ), 10% FBS ( k ) and 50% FBS ( l ). Scale bars are 100 nm ( b , j – l ) or 50 nm ( c – g ).

    Journal: Nature Communications

    Article Title: Dynamic protein coronas revealed as a modulator of silver nanoparticle sulphidation in vitro

    doi: 10.1038/ncomms11770

    Figure Lengend Snippet: Protein coronas modulate sulphide formation. Proposed mechanism of protein corona-modulated nano-Ag 2 S formation at Ag NPs, with hard corona proteins trapping Ag + released from the nanoparticle surface and soft corona proteins transporting said ions away from the sulphide-formation centres in the long-lived corona ( a ); TEM images of silver nanocubes after 24 h in RPMI-1640 cell culture medium supplemented with 1% FBS ( b ), followed by 6 days incubation in RPMI-1640 with 0% FBS (inset cartoon showing only hard corona around Ag NPs) ( c ), 1% FBS ( d ) or 10% FBS ( e ; common inset cartoon showing hard and soft coronas, as well as free bulk proteins around Ag NPs); TEM images of silver nanocubes after 7 days incubation in RPMI-1640 with 0.4 mg ml −1 BSA ( f ) or 4 mg ml −1 BSA ( g ) (common inset cartoon showing hard corona and free bulk proteins around Ag NPs); Ultraviolet–visible spectra of cubic ( h ) and quasi-spherical ( i ) Ag NPs after 24 h incubation in RPMI-1640 cell culture medium supplemented with 1, 10 or 50% FBS; TEM images of silver nanocubes after 24 h in RPMI-1640 supplemented with 1% FBS ( j ), 10% FBS ( k ) and 50% FBS ( l ). Scale bars are 100 nm ( b , j – l ) or 50 nm ( c – g ).

    Article Snippet: We incubated Ag NPs (cubic or quasi-spherical) in RPMI-1640 with or without added supplements of heat-inactivated FBS (HyClone; 1–50% by volume), for 1 or 7 days.

    Techniques: Transmission Electron Microscopy, Cell Culture, Incubation

    Silver sulphide forms close to the surface of Ag NPs. TEM image with arrows highlighting nano-Ag 2 S ( a , scale bar 50 nm), X-rays elemental mapping of Ag (red), S (blue, with white rings marking the approximate contour of the Ag NPs) and overlaid Ag and S ( b ), EDS spectrum—with arrows pointing at the peaks corresponding to each element—( c ) and diffraction pattern—arrow pointing at the diffraction line corresponding to monoclinic Ag 2 S—( d ) of silver nanocubes after 7 days incubation in RPMI-1640 supplemented with 1% FBS and formation of Ag 2 S at the surface of the Ag NPs.

    Journal: Nature Communications

    Article Title: Dynamic protein coronas revealed as a modulator of silver nanoparticle sulphidation in vitro

    doi: 10.1038/ncomms11770

    Figure Lengend Snippet: Silver sulphide forms close to the surface of Ag NPs. TEM image with arrows highlighting nano-Ag 2 S ( a , scale bar 50 nm), X-rays elemental mapping of Ag (red), S (blue, with white rings marking the approximate contour of the Ag NPs) and overlaid Ag and S ( b ), EDS spectrum—with arrows pointing at the peaks corresponding to each element—( c ) and diffraction pattern—arrow pointing at the diffraction line corresponding to monoclinic Ag 2 S—( d ) of silver nanocubes after 7 days incubation in RPMI-1640 supplemented with 1% FBS and formation of Ag 2 S at the surface of the Ag NPs.

    Article Snippet: We incubated Ag NPs (cubic or quasi-spherical) in RPMI-1640 with or without added supplements of heat-inactivated FBS (HyClone; 1–50% by volume), for 1 or 7 days.

    Techniques: Transmission Electron Microscopy, Incubation

    Silver nanoparticle dissolution is involved in nano-Ag 2 S formation. Ultraviolet–visible full spectra of quasi-spherical Ag NPs ( a ) and quadrupole peak detail of nanocubes ( b ) incubated (1 day: blue or 7 days: pink) in RPMI-1640 cell culture medium supplemented with 1% FBS, with or without added extra 10% (by mass) Ag + ions from AgNO 3 ; EDS spectra of the supernatant obtained after centrifugation of Ag NPs incubated for 7 days in RPMI-1640 with 1% FBS, before ( c ) and after ( d ) spiking with 5 nm PVP-coated Ag NPs, with dotted red line highlighting the presence of a silver signal only in the spiked sample.

    Journal: Nature Communications

    Article Title: Dynamic protein coronas revealed as a modulator of silver nanoparticle sulphidation in vitro

    doi: 10.1038/ncomms11770

    Figure Lengend Snippet: Silver nanoparticle dissolution is involved in nano-Ag 2 S formation. Ultraviolet–visible full spectra of quasi-spherical Ag NPs ( a ) and quadrupole peak detail of nanocubes ( b ) incubated (1 day: blue or 7 days: pink) in RPMI-1640 cell culture medium supplemented with 1% FBS, with or without added extra 10% (by mass) Ag + ions from AgNO 3 ; EDS spectra of the supernatant obtained after centrifugation of Ag NPs incubated for 7 days in RPMI-1640 with 1% FBS, before ( c ) and after ( d ) spiking with 5 nm PVP-coated Ag NPs, with dotted red line highlighting the presence of a silver signal only in the spiked sample.

    Article Snippet: We incubated Ag NPs (cubic or quasi-spherical) in RPMI-1640 with or without added supplements of heat-inactivated FBS (HyClone; 1–50% by volume), for 1 or 7 days.

    Techniques: Incubation, Cell Culture, Centrifugation

    Sulphur sources and the Ag:S ratio influence Ag 2 S formation. TEM images of Ag NPs after 7 days incubation in PBS ( a ), PBS supplemented with 1% FBS ( b ) and PBS supplemented with L -cysteine and L -methionine at the same concentrations of amino acids as those found in RPMI-1640 ( c ) and corresponding EDS spectra ( d ); TEM images and corresponding EDS spectra (insets) of Ag NPs after 7 days incubation in RPMI-1640 supplemented with 1% FBS, with initial silver concentrations of 2 μg ml −1 ( e ), 10 μg ml −1 (f) and 100 μg ml −1 ( g ), with elemental mapping images provided in Supplementary Fig. 26 . Scale bars are 100 nm ( a – c ) or 50 nm ( e – g ).

    Journal: Nature Communications

    Article Title: Dynamic protein coronas revealed as a modulator of silver nanoparticle sulphidation in vitro

    doi: 10.1038/ncomms11770

    Figure Lengend Snippet: Sulphur sources and the Ag:S ratio influence Ag 2 S formation. TEM images of Ag NPs after 7 days incubation in PBS ( a ), PBS supplemented with 1% FBS ( b ) and PBS supplemented with L -cysteine and L -methionine at the same concentrations of amino acids as those found in RPMI-1640 ( c ) and corresponding EDS spectra ( d ); TEM images and corresponding EDS spectra (insets) of Ag NPs after 7 days incubation in RPMI-1640 supplemented with 1% FBS, with initial silver concentrations of 2 μg ml −1 ( e ), 10 μg ml −1 (f) and 100 μg ml −1 ( g ), with elemental mapping images provided in Supplementary Fig. 26 . Scale bars are 100 nm ( a – c ) or 50 nm ( e – g ).

    Article Snippet: We incubated Ag NPs (cubic or quasi-spherical) in RPMI-1640 with or without added supplements of heat-inactivated FBS (HyClone; 1–50% by volume), for 1 or 7 days.

    Techniques: Transmission Electron Microscopy, Incubation

    Corona-mediated sulphidation of Ag NPs impacts particle toxicity. TEM images of partially sulphidated Ag NPs after pre-incubation in RPMI-1640 with 10% FBS ( a ) and completely sulphidated Ag NPs after pre-incubation in RPMI-1640 with 1% FBS ( b ); scale bars are 50 nm; Viability of J774 murine macrophages (as measured with MTT assays) after 24 h exposure to various concentrations (2, 5, 10, 15, 25, 50 and 100 μg ml −1 ) of Ag + ions (black diamonds), pristine Ag NPs (red triangles), partially sulphidated Ag NPs (blue squares) and completely sulphidated Ag NPs (orange circles); error bars are provided as standard deviation; statistically significant differences (two-tailed t -test, with all data sets showing normal distribution and similar variance values) as compared with the control are marked with ** P

    Journal: Nature Communications

    Article Title: Dynamic protein coronas revealed as a modulator of silver nanoparticle sulphidation in vitro

    doi: 10.1038/ncomms11770

    Figure Lengend Snippet: Corona-mediated sulphidation of Ag NPs impacts particle toxicity. TEM images of partially sulphidated Ag NPs after pre-incubation in RPMI-1640 with 10% FBS ( a ) and completely sulphidated Ag NPs after pre-incubation in RPMI-1640 with 1% FBS ( b ); scale bars are 50 nm; Viability of J774 murine macrophages (as measured with MTT assays) after 24 h exposure to various concentrations (2, 5, 10, 15, 25, 50 and 100 μg ml −1 ) of Ag + ions (black diamonds), pristine Ag NPs (red triangles), partially sulphidated Ag NPs (blue squares) and completely sulphidated Ag NPs (orange circles); error bars are provided as standard deviation; statistically significant differences (two-tailed t -test, with all data sets showing normal distribution and similar variance values) as compared with the control are marked with ** P

    Article Snippet: We incubated Ag NPs (cubic or quasi-spherical) in RPMI-1640 with or without added supplements of heat-inactivated FBS (HyClone; 1–50% by volume), for 1 or 7 days.

    Techniques: Transmission Electron Microscopy, Incubation, MTT Assay, Standard Deviation, Two Tailed Test

    Inflammatory gene expression profile in PGRN pretreated microglia. BV2 microglia were plated at 150,000 cells per well in a 12-well plate in DMEM/F-12 with 5% FBS. Cells were treated for 16 h with saline followed by 24 h of saline (Control), or saline

    Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience

    Article Title: Progranulin Does Not Bind Tumor Necrosis Factor (TNF) Receptors and Is Not a Direct Regulator of TNF-Dependent Signaling or Bioactivity in Immune or Neuronal Cells

    doi: 10.1523/JNEUROSCI.5336-12.2013

    Figure Lengend Snippet: Inflammatory gene expression profile in PGRN pretreated microglia. BV2 microglia were plated at 150,000 cells per well in a 12-well plate in DMEM/F-12 with 5% FBS. Cells were treated for 16 h with saline followed by 24 h of saline (Control), or saline

    Article Snippet: For recombinant PGRN co-addition experiments (see , ), bone marrow-derived macrophages (BMDMs) from C57BL/6 Grn +/+ or Grn +/− (heterozygous) male mice, generated as described previously , were harvested and plated at a density of 0.5 million cells per well in a 12-well tissue culture plate in CM, which consisted of DMEM/F12 (D6421; Sigma), 20% heat-inactivated FBS ( ; Atlanta Biologicals), 1% l -glutamine (Sigma), 1% penicillin/streptomycin (Sigma), and 20% CM from L929 (ATCC catalog #CCL1) mouse fibroblast cell line [DMEM (D5796, Invitrogen), 10% FBS (Sigma), 1% l -glutamine, 1% penicillin/streptomycin].

    Techniques: Expressing

    Activation of CD271 by nerve growth factor confers a more invasive phenotype in MOC2 cells and HSC3 cells by the induction of Snai2/Slug expression. (A-B) Comparison of cancer cell invasiveness in response to recombinant human nerve growth factor-β (rhNGF-β) treatment. Growth factor reduced Matrigel® Invasion Chambers were used to assess and compare the invasive potential of MOC2 vs. MOC2-CD271 cells (A), and that of HSC3 vs. HSC3-CD271 cells (B). Cells were cultured in low serum media (DMEM/F12 with 2% fetal bovine serum) for 24 hrs, and then, treated with rhNGF-β (100ng/mL). Cells were collected and incubated in invasion chambers with fibronectin (10ng/mL) as a chemoattractant for 48 hrs. Quantitation of invasion assay was performed by counting invaded cells in 6 random fields per chamber under light microscopy. Mean±SD is shown, **p

    Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

    Article Title: CD271 Confers an Invasive and Metastatic Phenotype of Head and Neck Squamous Cell Carcinoma Through the Upregulation of Slug

    doi: 10.1158/1078-0432.CCR-17-0866

    Figure Lengend Snippet: Activation of CD271 by nerve growth factor confers a more invasive phenotype in MOC2 cells and HSC3 cells by the induction of Snai2/Slug expression. (A-B) Comparison of cancer cell invasiveness in response to recombinant human nerve growth factor-β (rhNGF-β) treatment. Growth factor reduced Matrigel® Invasion Chambers were used to assess and compare the invasive potential of MOC2 vs. MOC2-CD271 cells (A), and that of HSC3 vs. HSC3-CD271 cells (B). Cells were cultured in low serum media (DMEM/F12 with 2% fetal bovine serum) for 24 hrs, and then, treated with rhNGF-β (100ng/mL). Cells were collected and incubated in invasion chambers with fibronectin (10ng/mL) as a chemoattractant for 48 hrs. Quantitation of invasion assay was performed by counting invaded cells in 6 random fields per chamber under light microscopy. Mean±SD is shown, **p

    Article Snippet: Cells were maintained in complete DMEM:F12 medium (DMEM:F12 1:1 with Glutamax (Gibco, Invitrogen, CA)) containing: 10% heat-inactivated FBS (Cellgro, MA), 100 IU/ml penicillin and 100 μg/ml streptomycin (Gibco, Invitrogen, CA).

    Techniques: Activation Assay, Expressing, Recombinant, Cell Culture, Incubation, Quantitation Assay, Invasion Assay, Light Microscopy

    Loss of GADD34 increased cellular damage and apoptosis in RAW 264.7 cells treated with LPS and Tyr/Cys-deprivation. (a). Knockdown of GADD34 by shRNA was evaluated. Real-time PCR analysis of GADD34 expression in GADD34-deficient (shGADD34) and control RAW 264.7 cells (shControl) after treatment with LPS and Tyr/Cys-deprivation for 8 h. Graph shows the relative expression as means ± SE of three independent experiments. (b). GADD34-deficient or control RAW 264.7 cells were treated with DMEM + 10%FBS (control) or LPS (1 μg/mL) in Tyr/Cys-deprivation medium for 8 h or 14 h. Cells were stained with 7-AAD and PE-labeled Annexin V and assessed by flow cytometry. (c). GADD34-deficient or control RAW 264.7 cells were treated with LPS with or without Tyr/Cys-deprivation for 14 h. Cells were stained with 7-AAD and PE-labeled Annexin V and assessed by flow cytometry. Relative % of Annexin V + 7AAD − apoptotic cells shown as means ± SE of three independent experiments. (d). BMDMs from GADD34 KO or WT were treated with LPS (1 μg/mL) combined with Tyr/Cys-deprivation for 8 h. Cells were stained with 7-AAD and PE-labeled Annexin V and assessed by flow cytometry. (e). GADD34-deficient or control RAW 264.7 cells were treated with LPS (+L; 1 μg/mL) and/or Tyr/Cys-deprivation for 24 h. Cell lysates were immunoblotted with anti-caspase 3 antibody and detected full length caspase 3 (35 kDa) and cleaved caspase 3 (17, 19 kDa). (f). GADD34-deficient or control RAW 264.7 cells were treated with DMEM + 10% FBS (control) or LPS (1 μg/mL) with or without Tyr/Cys-deprivation medium for 24 h. After treatment, cells were collected and counted by trypan-blue dye exclusion using a Burker-Turk cell count chamber. Data are means ± SE of cell number from three independent experiments. Data are representative of three independent experiments (b, d, e). * p

    Journal: Scientific Reports

    Article Title: GADD34 inhibits activation-induced apoptosis of macrophages through enhancement of autophagy

    doi: 10.1038/srep08327

    Figure Lengend Snippet: Loss of GADD34 increased cellular damage and apoptosis in RAW 264.7 cells treated with LPS and Tyr/Cys-deprivation. (a). Knockdown of GADD34 by shRNA was evaluated. Real-time PCR analysis of GADD34 expression in GADD34-deficient (shGADD34) and control RAW 264.7 cells (shControl) after treatment with LPS and Tyr/Cys-deprivation for 8 h. Graph shows the relative expression as means ± SE of three independent experiments. (b). GADD34-deficient or control RAW 264.7 cells were treated with DMEM + 10%FBS (control) or LPS (1 μg/mL) in Tyr/Cys-deprivation medium for 8 h or 14 h. Cells were stained with 7-AAD and PE-labeled Annexin V and assessed by flow cytometry. (c). GADD34-deficient or control RAW 264.7 cells were treated with LPS with or without Tyr/Cys-deprivation for 14 h. Cells were stained with 7-AAD and PE-labeled Annexin V and assessed by flow cytometry. Relative % of Annexin V + 7AAD − apoptotic cells shown as means ± SE of three independent experiments. (d). BMDMs from GADD34 KO or WT were treated with LPS (1 μg/mL) combined with Tyr/Cys-deprivation for 8 h. Cells were stained with 7-AAD and PE-labeled Annexin V and assessed by flow cytometry. (e). GADD34-deficient or control RAW 264.7 cells were treated with LPS (+L; 1 μg/mL) and/or Tyr/Cys-deprivation for 24 h. Cell lysates were immunoblotted with anti-caspase 3 antibody and detected full length caspase 3 (35 kDa) and cleaved caspase 3 (17, 19 kDa). (f). GADD34-deficient or control RAW 264.7 cells were treated with DMEM + 10% FBS (control) or LPS (1 μg/mL) with or without Tyr/Cys-deprivation medium for 24 h. After treatment, cells were collected and counted by trypan-blue dye exclusion using a Burker-Turk cell count chamber. Data are means ± SE of cell number from three independent experiments. Data are representative of three independent experiments (b, d, e). * p

    Article Snippet: RAW 264.7 cells were cultured in DMEM (Sigma, D5766) supplemented with 10% heat-inactivated FBS (Equitech-Bio Inc.).

    Techniques: shRNA, Real-time Polymerase Chain Reaction, Expressing, Staining, Labeling, Flow Cytometry, Cytometry, Cell Counting

    Expression of GADD34 induced by amino acid-deprivation and LPS. (a). Deprivation of grouped amino acids. RAW 264.7 cells were cultured with the indicated amino acid deprivation media for 4 h. Cell lysates were immunoblotted with anti-GADD34 and anti-β-actin antibodies. Β-actin was used as a control. Arrow indicates GADD34-specific bands. Asterisk indicates non-specific bands. Intensities of the bands were measured by densitometry. Graph shows the immunoblot relative intensities as means ± SE of 3 independent experiments. FBS; DMEM + 10% FBS, +15 A.A. (amino acid); Basal medium containing 15 amino acids ( Table 1 ) without FBS. Gly/Ser/Tyr/Cys deprivation; Arg/Gln/His deprivation; Ile/Leu/Lys/Met deprivation; Phe/Thr/Trp/Val deprivation. (b). Deprivation of single or two amino acids. Assay was performed as in a. (c). Effects of amino acid deprivation. Expression of GADD34 was analyzed for Leu/Lys, Arg or Trp deprivation compared to Tyr/Cys deprivation. (d). RAW 264.7 cells were stimulated with LPS (1 μg/mL) in DMEM + 10% FBS (control) or Tyr/Cys-deprivation medium for 4 h. GADD34 expression was determined by immunobloting. (e). Time course (0–24 h) of GADD34-expression in RAW 264.7 cells incubated in Tyr/Cys-deprivation medium with or without LPS (1 μg/mL). (f). BMDMs were stimulated with LPS (1 μg/mL) in control or Tyr/Cys- deprivation medium for 0–24 h. (g). Human macrophage THP1 cells were stimulated with LPS, deprived of Tyr/Cys or both for 8 h. The expression of GADD34 was determined by real-time PCR. Β-actin was used as an internal control. (h). GADD34 KO or WT mice were starved and injected with LPS (5 μg/g body weight). Expression of GADD34 in bone marrow was analyzed after 24 h starvation and LPS stimulation. The original immunoblots are presented in Supplementary Figure 3 . All immunoblots are representative of three independent experiments (a–f, h). Graph shows the relative expression as means ± SE of three independent experiments (a–d, g). * p

    Journal: Scientific Reports

    Article Title: GADD34 inhibits activation-induced apoptosis of macrophages through enhancement of autophagy

    doi: 10.1038/srep08327

    Figure Lengend Snippet: Expression of GADD34 induced by amino acid-deprivation and LPS. (a). Deprivation of grouped amino acids. RAW 264.7 cells were cultured with the indicated amino acid deprivation media for 4 h. Cell lysates were immunoblotted with anti-GADD34 and anti-β-actin antibodies. Β-actin was used as a control. Arrow indicates GADD34-specific bands. Asterisk indicates non-specific bands. Intensities of the bands were measured by densitometry. Graph shows the immunoblot relative intensities as means ± SE of 3 independent experiments. FBS; DMEM + 10% FBS, +15 A.A. (amino acid); Basal medium containing 15 amino acids ( Table 1 ) without FBS. Gly/Ser/Tyr/Cys deprivation; Arg/Gln/His deprivation; Ile/Leu/Lys/Met deprivation; Phe/Thr/Trp/Val deprivation. (b). Deprivation of single or two amino acids. Assay was performed as in a. (c). Effects of amino acid deprivation. Expression of GADD34 was analyzed for Leu/Lys, Arg or Trp deprivation compared to Tyr/Cys deprivation. (d). RAW 264.7 cells were stimulated with LPS (1 μg/mL) in DMEM + 10% FBS (control) or Tyr/Cys-deprivation medium for 4 h. GADD34 expression was determined by immunobloting. (e). Time course (0–24 h) of GADD34-expression in RAW 264.7 cells incubated in Tyr/Cys-deprivation medium with or without LPS (1 μg/mL). (f). BMDMs were stimulated with LPS (1 μg/mL) in control or Tyr/Cys- deprivation medium for 0–24 h. (g). Human macrophage THP1 cells were stimulated with LPS, deprived of Tyr/Cys or both for 8 h. The expression of GADD34 was determined by real-time PCR. Β-actin was used as an internal control. (h). GADD34 KO or WT mice were starved and injected with LPS (5 μg/g body weight). Expression of GADD34 in bone marrow was analyzed after 24 h starvation and LPS stimulation. The original immunoblots are presented in Supplementary Figure 3 . All immunoblots are representative of three independent experiments (a–f, h). Graph shows the relative expression as means ± SE of three independent experiments (a–d, g). * p

    Article Snippet: RAW 264.7 cells were cultured in DMEM (Sigma, D5766) supplemented with 10% heat-inactivated FBS (Equitech-Bio Inc.).

    Techniques: Expressing, Cell Culture, Western Blot, Incubation, Real-time Polymerase Chain Reaction, Mouse Assay, Injection

    QD232 inhibits cell migration. (A) A 24 h treatment with QD232 (1 μM) resulted in decreased migration of serum-starved MIA PaCa-2 cells through the membrane of a Boyden chamber in the presence of 10% FBS stimulation. Cells were imaged after fixing with methanol and staining with Giemsa using a Nikon microscope with 10× objective. (B) Quantification of data (means ± SD) shown in (A); effect induced by both concentrations of QD232 were significantly different from control (10% FBS only); P ). All concentrations of QD232, except the lowest (0.05μM in F, and 0.05 and 0.1 μM in G), had significant effects, compared with 10% FBS only; P

    Journal: British Journal of Pharmacology

    Article Title: Mechanisms underlying the cytotoxicity of a novel quinazolinedione-based redox modulator, QD232, in pancreatic cancer cells

    doi: 10.1111/bph.12855

    Figure Lengend Snippet: QD232 inhibits cell migration. (A) A 24 h treatment with QD232 (1 μM) resulted in decreased migration of serum-starved MIA PaCa-2 cells through the membrane of a Boyden chamber in the presence of 10% FBS stimulation. Cells were imaged after fixing with methanol and staining with Giemsa using a Nikon microscope with 10× objective. (B) Quantification of data (means ± SD) shown in (A); effect induced by both concentrations of QD232 were significantly different from control (10% FBS only); P ). All concentrations of QD232, except the lowest (0.05μM in F, and 0.05 and 0.1 μM in G), had significant effects, compared with 10% FBS only; P

    Article Snippet: Cell lines were maintained in the appropriate growth media [DMEM (Cellgro, Mediatech, Manassas, VA, USA) for PANC-1, MIA PaCa-2, and RPMI-1640 (Cellgro) for ASPC-1, BxPC-3 and HFF-1] containing 10% heat-inactivated FBS (Gemini-Bioproducts, West Sacramento, CA, USA) at 37°C in a humidified atmosphere of 5% CO2 .

    Techniques: Migration, Staining, Microscopy

    Effect of ST258 KP strains on inflammatory cytokines production by monocytes and MDC. Monocytes or MDC were cultured at 10 6 cells/ml in RPMI with 10% FBS in the presence or absence (US) of UV-inactivated ST258-KP strains at 1:10 ratio or of LPS (400 ng/ml). Conditioned media were collected after 16 hours of culture. Cytokine production was measured by Immunoplex array. Box-chart plots show cytokine production from 6 different healthy donors. The boxes extend from SE; the horizontal line represents the median, asterisks indicate the mean value. Statistical analysis was performed by Student’s t-test and p ≤ 0.05 was considered significant.

    Journal: PLoS ONE

    Article Title: Differences in Inflammatory Response Induced by Two Representatives of Clades of the Pandemic ST258 Klebsiella pneumoniae Clonal Lineage Producing KPC-Type Carbapenemases

    doi: 10.1371/journal.pone.0170125

    Figure Lengend Snippet: Effect of ST258 KP strains on inflammatory cytokines production by monocytes and MDC. Monocytes or MDC were cultured at 10 6 cells/ml in RPMI with 10% FBS in the presence or absence (US) of UV-inactivated ST258-KP strains at 1:10 ratio or of LPS (400 ng/ml). Conditioned media were collected after 16 hours of culture. Cytokine production was measured by Immunoplex array. Box-chart plots show cytokine production from 6 different healthy donors. The boxes extend from SE; the horizontal line represents the median, asterisks indicate the mean value. Statistical analysis was performed by Student’s t-test and p ≤ 0.05 was considered significant.

    Article Snippet: RPMI 1640, antibiotics (penicillin/streptomycin), L-glutamine, heat-inactivated FBS were purchased from Euroclone (Pero, Italy) and used for cultures with dendritic cells.

    Techniques: Cell Culture

    Effect of ST258 KP strains on NLRP3 expression. Panel A: Effect of ST258 KP strains on NLRP3 protein expression. Monocytes or MDC from 3 different donors were cultured at 10 6 cells/ml in RPMI containing 10% FBS with live ST258 KP- strains at 1:1 ratio for 7 hours. At the end of incubation cells were lysed and analyzed by Western Blot. Results from one representative experiment are shown. The histogram below shows the results of densitometric analysis from the three different experiments (mean ± SE). Panel B: Effect of ST258 KP strains on NLRP3 and pro- IL-1 β (gene) expression. Monocytes or MDC were cultured at 10 6 cells/ml in the presence or absence of live ST258 KP strains for 4 hours at 1:1 cell ratio. Cells were lysed to obtain total RNA. The NLRP3 and pro- IL-1 β gene expression was evaluated by RT-PCR using specific primers. Results are expressed as mean ± SE of mRNA relative amounts (2 -ΔCT ) of experimental triplicates. Histograms show one representative experiment out of four performed. Statistical analysis was performed by Student t -test and p ≤ 0.05 was considered significant.

    Journal: PLoS ONE

    Article Title: Differences in Inflammatory Response Induced by Two Representatives of Clades of the Pandemic ST258 Klebsiella pneumoniae Clonal Lineage Producing KPC-Type Carbapenemases

    doi: 10.1371/journal.pone.0170125

    Figure Lengend Snippet: Effect of ST258 KP strains on NLRP3 expression. Panel A: Effect of ST258 KP strains on NLRP3 protein expression. Monocytes or MDC from 3 different donors were cultured at 10 6 cells/ml in RPMI containing 10% FBS with live ST258 KP- strains at 1:1 ratio for 7 hours. At the end of incubation cells were lysed and analyzed by Western Blot. Results from one representative experiment are shown. The histogram below shows the results of densitometric analysis from the three different experiments (mean ± SE). Panel B: Effect of ST258 KP strains on NLRP3 and pro- IL-1 β (gene) expression. Monocytes or MDC were cultured at 10 6 cells/ml in the presence or absence of live ST258 KP strains for 4 hours at 1:1 cell ratio. Cells were lysed to obtain total RNA. The NLRP3 and pro- IL-1 β gene expression was evaluated by RT-PCR using specific primers. Results are expressed as mean ± SE of mRNA relative amounts (2 -ΔCT ) of experimental triplicates. Histograms show one representative experiment out of four performed. Statistical analysis was performed by Student t -test and p ≤ 0.05 was considered significant.

    Article Snippet: RPMI 1640, antibiotics (penicillin/streptomycin), L-glutamine, heat-inactivated FBS were purchased from Euroclone (Pero, Italy) and used for cultures with dendritic cells.

    Techniques: Expressing, Cell Culture, Incubation, Western Blot, Reverse Transcription Polymerase Chain Reaction

    CPX does not alter mRNA expression, but inhibits protein synthesis and promotes protein degradation of VEGFR-3 A, CPX did not affect VEGFR-3 mRNA level. Total RNA was extracted from LECs treated with CPX (0-5 μM) for 24 h (Left panel) or with CPX (5 μM) for 0-24 h (Right panel), followed by semi-quantitative RT-PCR. β-actin was used as a loading control. B, CPX inhibited protein synthesis of VEGFR-3 in LECs. LECs were pretreated with CPX (0-5 μM) for 24 h (Left panel) or with CPX (5 μM) for 0-24 h (Right panel), and then pulsed with 35 S-Met/Cys for 4 h, followed by immunoprecipitation with antibodies to VEGFR-3. The immunoprecipitates were separated by SDS-PAGE and transferred to PVDF membranes, followed by autoradiography. GAPDH served as an internal control. C, CPX promoted protein degradation of VEGFR-3 in LECs. LECs, grown in 10% FBS-DMEM medium, were exposed to cycloheximide (CHX, 50 μg/ml), in the presence or absence of CPX (5 μM), for 0-12 h, followed by Western blot analysis with indicated antibodies.

    Journal: Oncogene

    Article Title: The fungicide ciclopirox inhibits lymphatic endothelial cell tube formation by suppressing VEGFR-3-mediated ERK signaling pathway

    doi: 10.1038/onc.2010.590

    Figure Lengend Snippet: CPX does not alter mRNA expression, but inhibits protein synthesis and promotes protein degradation of VEGFR-3 A, CPX did not affect VEGFR-3 mRNA level. Total RNA was extracted from LECs treated with CPX (0-5 μM) for 24 h (Left panel) or with CPX (5 μM) for 0-24 h (Right panel), followed by semi-quantitative RT-PCR. β-actin was used as a loading control. B, CPX inhibited protein synthesis of VEGFR-3 in LECs. LECs were pretreated with CPX (0-5 μM) for 24 h (Left panel) or with CPX (5 μM) for 0-24 h (Right panel), and then pulsed with 35 S-Met/Cys for 4 h, followed by immunoprecipitation with antibodies to VEGFR-3. The immunoprecipitates were separated by SDS-PAGE and transferred to PVDF membranes, followed by autoradiography. GAPDH served as an internal control. C, CPX promoted protein degradation of VEGFR-3 in LECs. LECs, grown in 10% FBS-DMEM medium, were exposed to cycloheximide (CHX, 50 μg/ml), in the presence or absence of CPX (5 μM), for 0-12 h, followed by Western blot analysis with indicated antibodies.

    Article Snippet: Human embryonic kidney (HEK) 293 (American Type Culture Collection, Manassas, VA), 293TD and 293A cells (Invitrogen, Carlsbad, CA, USA) were grown in antibiotic-free Dulbecco’s modified Eagle medium (DMEM) (Mediatech) supplemented with 10% heat-inactivated FBS and non-essential amino acid (Mediatech) at 37°C and 5% CO2 .

    Techniques: Expressing, Quantitative RT-PCR, Immunoprecipitation, SDS Page, Autoradiography, Western Blot