cd31/pecam-1 antibody - bsa free  (Bio-Techne corporation)

Journal: Nature Communications

Article Title: Integrin α3β1 promotes vessel formation of glioblastoma-associated endothelial cells through calcium-mediated macropinocytosis and lysosomal exocytosis

doi: 10.1038/s41467-022-31981-2

Figure Lengend Snippet: a , b Total RNA was extracted from 3 NECs (376, 422, and 623) and 3 TECs (isolates cw880, ccf2352, and ccf2390) when similar densities of tubes had formed on Matrigel, and c , d from monolayers of 6 TECs (cw850, cw880, ccf2277, ccf2352, ccf2388, and ccf2390) and 3 NECs (375, 422, and 623), followed by gene expression microarray (HG-U219). a , c Genes upregulated over 2-fold in TECs compared to NECs were subject to gene set enrichment and pathway analysis using DAVID Bioinformatics Resources 6.8 ( https://david.ncifcrf.gov ), where p -values were calculated by Fisher’s exact test. b , d Expression profiles of integrins and genes relevant to angiogenesis both on Matrigel and in monolayer are shown as heatmaps; p -values were obtained by two-sided exact-Wilcoxon-rank-sum tests, without adjustment for multiple comparisons. e Cell lysates derived from the same experiment with monolayers of NECs (623, 376, and 422) and TECs (ccf2445, ccf2566, and ccf2515) were subjected to western blot analysis with the indicated antibodies in parallel. f Representative confocal images of integrin α3 staining of tubes formed by NECs (376) or TECs (ccf2566) plated on Matrigel (24 h). Graph represents the integrated densities of integrin α3-immunofluorescence. g Paraffin sections of human GBM and normal brain (NB) samples were double-labeled for integrin α3 and vwf, followed by DAPI staining. Representative images shown. Colocalization of the integrin α3 signal with the vwf signal for GBM-sections was 86%, based on the Manders’ Coefficient (M1 = 0.863; M2 = 0.515). h Human GBM tissue arrays containing NB were double-labeled for vwf and integrin α3, CD151, or integrin α6. Statistical analysis: two-sided Wilcoxon-rank-sum-tests; red lines denote means and black lines denote 95% confidence intervals. i Tumor sections from mouse glioma were double-labeled by immunofluorescence for integrin α3, CD151, or integrin α6 and CD31. Average fluorescence intensities of integrin α3, CD151, or integrin α6 on CD31-positive pixels/field were obtained from tumor or adjacent NB using ImageJ and represented as boxplots. Boxes indicate first and third quartiles, bands indicate medians, and whiskers indicate ±1.5 interquartile range. Statistical analysis: linear mixed model; red bars, means. ( h , i ) On the x axis, n = number of different fields. e – g Independent experiments were performed at least two times with similar results. Source data are provided as a source file.

Article Snippet: The primary antibodies used for immunofluorescence analysis of human cells and tissues were purchased as follows: Rabbit anti-CD31 antibody (NBP1-71663) from Novus Biologicals; mouse monoclonal anti-integrin α3 subunit (MAB1952z, clone P1B5); rabbit anti-vwf (AB7356) from Millipore; rabbit anti-VE-Cadherin antibody (ab33168), rabbit anti-integrin α3 subunit (ab131055), rabbit anti-CD151 (ab201174), mouse anti-vwf (ab68545), rabbit anti-SNX5 antibody (ab180520, clone EPR14368) and rabbit anti-LAMP1 antibody (ab24170) from Abcam; rabbit anti-integrin α6 subunit (T0919) from Epitomics; goat anti-laminin α5 IgG (sc-16592) from Santa Cruz, and mouse anti-LAMP1 antibody (H4A3) from the Developmental Studies Hybridoma Bank.

Techniques: Gene Expression, Microarray, Expressing, Derivative Assay, Western Blot, Staining, Immunofluorescence, Labeling, Fluorescence

Journal: Nature Communications

Article Title: Integrin α3β1 promotes vessel formation of glioblastoma-associated endothelial cells through calcium-mediated macropinocytosis and lysosomal exocytosis

doi: 10.1038/s41467-022-31981-2

Figure Lengend Snippet: a Schematic diagram of the experimental procedure. Briefly, GBM tumors (ccf4259, ccf4268, ccf4272, and DI247) cut into ~4 mm in diameter were cultured in Matrigel in neurobasal medium supplemented with 1 ng/ml EGF and 1 ng/ml bFGF. On day 2, anti-integrin α3 subunit blocking antibody or control mouse IgG was injected at three places per tumor piece to achieve ~5 µg/ml in tumor volume. The same injection was repeated on day 6. On day 9, tumors were fixed in 10% formalin and paraffin-embedded. Sections were labeled with rabbit anti-CD31 antibody and Alexa-Fluor-488 goat anti-rabbit IgG, followed by DAPI nuclear stain and imaging. b Representative immunofluorescent images from organotypic culture of tumor ccf4272 are shown. c Percent CD31-positive area per field was obtained by image processing using ImageJ. n denotes the number of different fields, for ccf4269, n = 44 for mouse IgG and n = 95 for anti-integrin α3; for ccf4268, n = 39 for mouse IgG and n = 103 for anti-integrin α3; for ccf4272, n = 57 for mouse IgG and n = 82 for anti-integrin α3; for DI-247, n = 59 for mouse IgG and n = 88 for anti-integrin α3. The boxes indicate the first and third quartiles, bands indicate medians, and the whiskers indicate ±1.5 interquartile range. A linear mixed model was used for statistical analysis ( p < 0.001). Source data are provided as a source data file.

Article Snippet: The primary antibodies used for immunofluorescence analysis of human cells and tissues were purchased as follows: Rabbit anti-CD31 antibody (NBP1-71663) from Novus Biologicals; mouse monoclonal anti-integrin α3 subunit (MAB1952z, clone P1B5); rabbit anti-vwf (AB7356) from Millipore; rabbit anti-VE-Cadherin antibody (ab33168), rabbit anti-integrin α3 subunit (ab131055), rabbit anti-CD151 (ab201174), mouse anti-vwf (ab68545), rabbit anti-SNX5 antibody (ab180520, clone EPR14368) and rabbit anti-LAMP1 antibody (ab24170) from Abcam; rabbit anti-integrin α6 subunit (T0919) from Epitomics; goat anti-laminin α5 IgG (sc-16592) from Santa Cruz, and mouse anti-LAMP1 antibody (H4A3) from the Developmental Studies Hybridoma Bank.

Techniques: Cell Culture, Blocking Assay, Control, Injection, Labeling, Staining, Imaging