vitronectin primary antibody  (R&D Systems)

Journal: PLoS ONE

Article Title: Functional role of vitronectin in breast cancer

doi: 10.1371/journal.pone.0242141

Figure Lengend Snippet: Displays mean fold change and standard error (SE) from the RPPA dataset, comparing race and tumor subtypes to the control group. Data allows us to compare level of vitronectin in serum. The mean fold change is relative to the control population for the same ethnicity.

Article Snippet: The vitronectin primary antibody (RD systems, MAB2349), PI3K (Cell Signaling Technology, 4292), P-AKT (Cell Signaling Technology, #9271), AKT (Cell Signaling Technology, #9272) were diluted with antibody diluent (ProteinSimple).

Techniques: Control

Journal: PLoS ONE

Article Title: Functional role of vitronectin in breast cancer

doi: 10.1371/journal.pone.0242141

Figure Lengend Snippet: A . Image displays the RPPA dot blot which was stained with the vitronectin antibody. Intensity values were taken from here and then analyzed. B-F . Displays comparisons of mean relative values of each group using standard error for error bars. B . analyzes overall serum vitronectin levels in control group (n = 40) and cancer group (n = 200). C . analyzes serum vitronectin levels (overall) in control groups and compared with the other tumor subtypes. Her2+ and LB2 show significant differences when compared to control group. D . compares control and cancer groups by ethnicity, and we find that CA cancer shows a significantly higher vitronectin concentration levels. E . we see that it compares vitronectin levels of AA and WA patients by dividing into various tumor characteristics. However, we found that there are significant differences in Her2+ and LB2 subtypes in the AA group when compared to AA controls. F . compares recurrent versus non-recurrent patient samples by ethnic groups. We found that there is a significant difference in serum vitronectin levels in non-recurrent CA compared to the control and recurrent groups, while AA samples had similar levels of serum vitronectin in all three groups.

Article Snippet: The vitronectin primary antibody (RD systems, MAB2349), PI3K (Cell Signaling Technology, 4292), P-AKT (Cell Signaling Technology, #9271), AKT (Cell Signaling Technology, #9272) were diluted with antibody diluent (ProteinSimple).

Techniques: Dot Blot, Staining, Control, Concentration Assay

Journal: PLoS ONE

Article Title: Functional role of vitronectin in breast cancer

doi: 10.1371/journal.pone.0242141

Figure Lengend Snippet: The data above is validation of data shown in , that showed potential racial disparities in tumor subtypes in vitronectin concentration levels of serum. A . Displays the relative concentration levels of vitronectin in AA Her2+ patients in serum compared with corresponding control samples. B . Relative serum concentration levels of vitronectin in WA Her2+ patients and compared with healthy patients C . Displays the relative serum concentration levels of vitronectin in AA LB2 patients and control serum. D . Displays the relative concentration levels of vitronectin in CA LB2 patients in serum.

Article Snippet: The vitronectin primary antibody (RD systems, MAB2349), PI3K (Cell Signaling Technology, 4292), P-AKT (Cell Signaling Technology, #9271), AKT (Cell Signaling Technology, #9272) were diluted with antibody diluent (ProteinSimple).

Techniques: Biomarker Discovery, Concentration Assay, Control

Journal: PLoS ONE

Article Title: Functional role of vitronectin in breast cancer

doi: 10.1371/journal.pone.0242141

Figure Lengend Snippet: A . Overall concentration level of vitronectin in 21 recurrence samples (n = 21) and compared serum vitronectin concentration levels to 21 non-recurrence samples of same IHC sub-groups. B . Receiver Operation Characteristic (ROC) Curve and, the statistical evaluation of vitronectin concentration in serum samples between non-recurrence vs recurrence patients. We tested to see if vitronectin could be used as a recurrence biomarkers and, we found that the area under curve (AUC) is 0.7107 with a p-value 0.0940.

Article Snippet: The vitronectin primary antibody (RD systems, MAB2349), PI3K (Cell Signaling Technology, 4292), P-AKT (Cell Signaling Technology, #9271), AKT (Cell Signaling Technology, #9272) were diluted with antibody diluent (ProteinSimple).

Techniques: Concentration Assay

Journal: PLoS ONE

Article Title: Functional role of vitronectin in breast cancer

doi: 10.1371/journal.pone.0242141

Figure Lengend Snippet: A . Displays the pseudo blot extracted from SimpleWestern experiments. There are four cell-lines- MDA-MB-231, MCF7, MDA-MB-468, HCC1599, were used to evaluate the signaling cascade relationship. Different protein expression levels were normalized with an averaged house-keeping GAPDH and β-actin expression B . Compares vitronectin concentration levels in four BC cell lines. C . PI3K concentration levels in BC same cell-lines. D . compares AKT concentration and, E . P-AKT concentration levels in the same four BC cell lines.

Article Snippet: The vitronectin primary antibody (RD systems, MAB2349), PI3K (Cell Signaling Technology, 4292), P-AKT (Cell Signaling Technology, #9271), AKT (Cell Signaling Technology, #9272) were diluted with antibody diluent (ProteinSimple).

Techniques: Expressing, Concentration Assay

Journal: Applied Bionics and Biomechanics

Article Title: Fluid Shear Stress and Fibroblast Growth Factor-2 Increase Endothelial Cell-Associated Vitronectin

doi: 10.1155/2017/9040161

Figure Lengend Snippet: Both shear stress and FGF-2 increased cell-associated vitronectin. HUVECs were exposed to steady laminar flow (20 dynes/cm 2 ) or oscillating disturbed flow (4 ± 6 dynes/cm 2 shear stress, 1 Hz) in a cone and plate device, or 50 ng/ml FGF-2 for 24 hours. Samples were then analyzed for vitronectin by Western blot. (a) HUVEC phase contrast microscopy images following 24 hours of flow or FGF-2 (10x). (b) Vitronectin (VTN) Western blot (one sample out of three replicates), with GAPDH loading control and quantification of the 75/65 kDa band (normalized to GAPDH). ∗ p < 0.05, ∗∗ p < 0.01. One representative experiment of two.

Article Snippet: Membranes were incubated overnight at 4°C with vitronectin primary antibody (sc-15332, Santa Cruz), followed by an anti-rabbit secondary horseradish peroxidase-conjugated antibody (Promega) for 2 hours at room temperature.

Techniques: Shear, Western Blot, Microscopy, Control

Journal: Applied Bionics and Biomechanics

Article Title: Fluid Shear Stress and Fibroblast Growth Factor-2 Increase Endothelial Cell-Associated Vitronectin

doi: 10.1155/2017/9040161

Figure Lengend Snippet: FGF-2 increased cell-associated vitronectin in (a) HUVEC (human umbilical vein endothelial cells), (b) BBmVEC (bovine brain microvascular endothelial cells), (c) PAEC (porcine aortic endothelial cells), and (d) PSMC (porcine vascular smooth muscle cells). Cells seeded on native collagen-coated substrates were stimulated with 50 ng/ml FGF-2 for 24, 48, and 72 hours. Cell extracts were collected, and normalized protein samples were analyzed by Western blot. VTN intensity was quantified and normalized to GAPDH or β -actin and then normalized to untreated cell VTN levels at 24 hours. # p < 0.05, ∗ p < 0.01, and ∗∗ p < 0.001 compared to those of the untreated cells. n = 3, one representative experiment of two.

Article Snippet: Membranes were incubated overnight at 4°C with vitronectin primary antibody (sc-15332, Santa Cruz), followed by an anti-rabbit secondary horseradish peroxidase-conjugated antibody (Promega) for 2 hours at room temperature.

Techniques: Western Blot

Journal: Applied Bionics and Biomechanics

Article Title: Fluid Shear Stress and Fibroblast Growth Factor-2 Increase Endothelial Cell-Associated Vitronectin

doi: 10.1155/2017/9040161

Figure Lengend Snippet: HUVEC produced vitronectin mRNA, but the FGF-2-induced increased in cell-associated vitronectin which came from the serum. (a) mRNA was isolated via reverse transcriptase PCR in HUVEC which were treated with 50 ng/ml FGF-2 for 24 hours. (b) Cell-associated vitronectin was analyzed by Western blot in HUVEC treated with 50 ng/ml FGF-2 for 24 hours in EBM-2 0% FBS, 1 μ g/ml mVTN, or 5% FBS. Vitronectin band intensity was quantified and normalized to GAPDH and then normalized to vitronectin levels in untreated cells in the serum-free medium. (c) Plasmin enzymatic activity was analyzed in HUVEC treated with 50 ng/ml FGF-2 for 24 hours in EBM-2 0% FBS, 1 μ g/ml mVTN, or 5% FBS by the Chromozym assay. All samples were normalized to untreated cells in the serum-free medium. # p < 0.05 and ∗∗ p < 0.001 compared to those of the untreated cells in the serum-free medium. n = 3, one representative experiment of three.

Article Snippet: Membranes were incubated overnight at 4°C with vitronectin primary antibody (sc-15332, Santa Cruz), followed by an anti-rabbit secondary horseradish peroxidase-conjugated antibody (Promega) for 2 hours at room temperature.

Techniques: Produced, Isolation, Reverse Transcription, Western Blot, Activity Assay

Journal: Applied Bionics and Biomechanics

Article Title: Fluid Shear Stress and Fibroblast Growth Factor-2 Increase Endothelial Cell-Associated Vitronectin

doi: 10.1155/2017/9040161

Figure Lengend Snippet: FGF-2 stimulation increased basal and intracellular vitronectin. Confocal microscopy images of cells labeled for vitronectin (AF488, green) and nuclei with (Hoescht, blue). HUVEC treated with 50 ng/ml FGF-2 for 24 hours were permeabilized with Triton X-100 to label membrane and cytoplasmic vitronectin. Samples were imaged in a z-stack (1 μ m depth, 12 images per sample) to obtain fluorescence intensity throughout the cell. Images in the z-stack were classified as basal (below the nucleus), intracellular (at the same levels as the nucleus), and apical (above the nucleus). Mean intensity values were quantified with ImageJ. All samples were normalized to total vitronectin in untreated cells. # p < 0.05 versus that of the untreated cells. n = 3, one representative experiment of three.

Article Snippet: Membranes were incubated overnight at 4°C with vitronectin primary antibody (sc-15332, Santa Cruz), followed by an anti-rabbit secondary horseradish peroxidase-conjugated antibody (Promega) for 2 hours at room temperature.

Techniques: Confocal Microscopy, Labeling, Membrane, Fluorescence

Journal: Applied Bionics and Biomechanics

Article Title: Fluid Shear Stress and Fibroblast Growth Factor-2 Increase Endothelial Cell-Associated Vitronectin

doi: 10.1155/2017/9040161

Figure Lengend Snippet: FGF-2-induced cell-associated vitronectin and plasminogen system activity were abrogated by genistein, a protein tyrosine kinase inhibitor. HUVEC were exposed to 30 μ g/ml genistein for 2 hours prior to stimulation with 50 ng/ml FGF-2 for 24 hours. (a) Samples were fixed and permeabilized with Triton X-100 to visualize total vitronectin. Samples were then labeled for vitronectin (green) and nuclei (blue). Samples were imaged in a z-stack (1 μ M depth, 12 images per sample) to obtain fluorescence intensity throughout the cell. Mean intensity values were quantified with ImageJ. (b) Cell-associated vitronectin was analyzed by Western blot in cells treated with FGF-2 and genistein. Vitronectin band intensity was quantified and normalized to GAPDH and then normalized to untreated cell levels. (c) Plasmin enzymatic activity was determined using the Chromozym assay in HUVEC treated with FGF-2 and genistein. Samples were normalized to untreated cells. # p < 0.05 and ∗ p < 0.01 compared to those of the untreated cells. n = 3, one representative experiment of two.

Article Snippet: Membranes were incubated overnight at 4°C with vitronectin primary antibody (sc-15332, Santa Cruz), followed by an anti-rabbit secondary horseradish peroxidase-conjugated antibody (Promega) for 2 hours at room temperature.

Techniques: Activity Assay, Labeling, Fluorescence, Western Blot

Journal: Applied Bionics and Biomechanics

Article Title: Fluid Shear Stress and Fibroblast Growth Factor-2 Increase Endothelial Cell-Associated Vitronectin

doi: 10.1155/2017/9040161

Figure Lengend Snippet: FGF-2-induced cell-associated vitronectin and plasminogen system activity were abrogated when the α v β 5 integrin was blocked. HUVEC were treated with 1 μ g/ml anti- α v β 5 -blocking antibody concurrently with 50 ng/ml FGF-2 for 24 hours. (a) Samples were fixed and permeabilized with Triton X-100 to visualize total vitronectin. Samples were then labeled for vitronectin (green) and nuclei (blue). Samples were imaged in a z-stack (1 μ M depth, 12 images per sample) to obtain fluorescence intensity throughout the cell. Mean intensity values were quantified with ImageJ. (b) Cell-associated vitronectin was analyzed by Western blot in cells treated with FGF-2 and genistein. Vitronectin band intensity was quantified and normalized to GAPDH and then normalized to untreated cell levels. (c) Plasmin enzymatic activity was determined using the Chromozym assay in HUVEC treated with FGF-2 and genistein. Samples were normalized to untreated cells. # p < 0.05 and ∗ p < 0.01 compared to those of the untreated cells. n = 3, one representative experiment of two.

Article Snippet: Membranes were incubated overnight at 4°C with vitronectin primary antibody (sc-15332, Santa Cruz), followed by an anti-rabbit secondary horseradish peroxidase-conjugated antibody (Promega) for 2 hours at room temperature.

Techniques: Activity Assay, Blocking Assay, Labeling, Fluorescence, Western Blot

Journal: Applied Bionics and Biomechanics

Article Title: Fluid Shear Stress and Fibroblast Growth Factor-2 Increase Endothelial Cell-Associated Vitronectin

doi: 10.1155/2017/9040161

Figure Lengend Snippet: FGF-2-induced cell-associated vitronectin and plasminogen system activity were abrogated by silencing the β 5 integrin with siRNA. HUVEC were transfected with β 5 siRNA for 72 hours. Samples were then treated with 50 ng/ml FGF-2 for 24 hours. (a) Samples were fixed and permeabilized with Triton X-100 to visualize total vitronectin. Samples were then labeled for vitronectin (green) and nuclei (blue). Samples were imaged in a z-stack (1 μ M depth, 12 images per sample) to obtain fluorescence intensity throughout the cell. Mean intensity values were quantified with ImageJ. (b) Cell-associated vitronectin was analyzed by Western blot in cells treated with FGF-2 and genistein. Vitronectin band intensity was quantified and normalized to GAPDH and then normalized to untreated cell levels. (c) Plasmin enzymatic activity was determined using the Chromozym assay in HUVEC treated with FGF-2 and genistein. Samples were normalized to untreated cells. # p < 0.05 and ∗ p < 0.01 compared to those of the untreated cells. n = 3, one representative experiment of two.

Article Snippet: Membranes were incubated overnight at 4°C with vitronectin primary antibody (sc-15332, Santa Cruz), followed by an anti-rabbit secondary horseradish peroxidase-conjugated antibody (Promega) for 2 hours at room temperature.

Techniques: Activity Assay, Transfection, Labeling, Fluorescence, Western Blot