Human Survivin Antibody

Detection of Survivin in Human PBMC lymphocytes by Flow Cytometry. Human peripheral blood mononuclear cells were labeled with Human Survivin Monoclonal Antibody (Catalog # MAB886, filled histogram) or isotype control antibody (Catalog # MAB002, open histogram), followed by Allophycocyanin-conjugated Anti-Mouse IgG F(ab')₂Secondary Antibody (Catalog # F0101B). To facilitate intracellular staining, cells were fixed with PFA and permeabilized with saponin.

Detection of Survivin by Flow Cytometry Exposure to survivin enriched PD-1+ Bcl-6+ subset of Tfh via STAT3 dependent mechanismsDBA/1 mice were immunized with survivin-derived peptide (100 μg/mouse × 4, subcutaneously). Control mice were immunized with ovalbumin-derived peptide (OVA). Both groups were then subjected to collagen-induced arthritis. Anti-survivin IgG antibodies, anti-Fcg antibodies, & survivin levels in serum were measured by ELISA A. Flow cytometry analysis of the expression of survivin B. in CD44hiCD4+ lymphocytes, & expression of PD-1 C. & Bcl-6 E. in survivin+CXCR5+ CD4+ lymphocytes in spleen (SPL) & lymph nodes (LN). PD-1 expression correlated to the size of CXCR5+survivin+ population D. Dots represent individual mice & the horizontal line shows median of the group. Protein levels of active STAT3 phosphorylated at Y705 (pStat3), total Stat3, Bcl-6 & actin in spleen were analyzed by the Western blot F. The levels of each protein were quantified in ratio to actin of each sample. Quantification of the detected bands is presented as box plots. Transcription of Bcl-6, cMaf & IL-21 in the spleen was analyzed by RT-PCR & presented in relative quantity (RQ) to the median of the control group G. Anti-survivin antibodies in serum correlated with the size of survivin+ CD4 population in spleen of survivin-immunized mice H. Comparison of the correlations in the survivin- & OVA-immunized groups is done by the Fisher r-to-z transformation analysis. Anti-Fcg antibodies correlated with the intensity of PD-1 on CXCR5+survivin+ CD4 lymphocytes I., as measured by flow cytometry. Box plots with line represent IQR of the group & median, respectively, & error lines indicate min & max values. The Mann-Whitney U-test was used to compare differences between groups. Correlation analyses were performed using Spearman's test. Image collected & cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/26343374), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Survivin by Flow Cytometry Survivin positive subset of CD44hi CD4 lymphocytes in mouse possess a complete phenotype of Tfh cells Spleen and lymph nodes from collagen II immunized arthritic (CIA) mice were analyzed for expression of survivin and Bcl-6 using flow cytometry A. Cells were gated on memory CD44hiCD4+ lymphocytes. Expression of CXCR5 B. and PD-1 C. was investigated within Bcl-6+ survivin+ and Bcl-6− survivin+ cells. Dots represent individual mice and the horizontal line shows median of the group. Survivin translation in CIA mice was inhibited by shRNA-producing constructs provided as a single intra-peritoneal injection (shSurv16, n = 10, or shSurv13+16, n = 10). Control mice were treated with a non-targeting RNA construct (shNT, n = 9). Survivin expression in spleen was analyzed by flow cytometry 12 days after the injection. Cells were gated on CD4+ lymphocytes. Intensity of survivin expression (MFI) within the groups is shown by a representative histogram and summarized in a box plot D. Survivin expression (MFI) on CD4 lymphocytes correlated to the size of CD44hiCD62L+ population E. Expression of CXCR5 on CD44hiCD4+ lymphocytes in the groups is shown as box plot F. CXCR5+ population correlates with the intensity of survivin G. and with Bcl-6 mRNA H. Boxes and lines represent IQR and median, respectively, and error lines indicate min and max values. The Mann-Whitney U-test was used to compare differences between groups. Correlation analyses were performed using Spearman's test. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/26343374), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Survivin by Flow Cytometry Survivin expression is an essential feature of human CXCR5+ Tfh cell phenotypeIntracellular expression of survivin was investigated in memory (CD45RA−) or naïve (CD45RA+) CD4+ T cells of RA patients (n = 21) and healthy controls (n = 10) using flow cytometry. Cells are gated on CD4+ lymphocytes. Box plots show the frequency of survivin+ cells A. and the mean fluorescence intensity (MFI) of survivin B. Expression of CXCR5 C. within survivin+ and survivin− CD4+ cells, and Bcl-6 D. within survivin+ and survivin− memory (CD45RA−) CD4+ cells of RA patients. The intensity of survivin expression E. within Bcl-6+ and Bcl-6− survivin+ CXCR5+ CD4 cells. The Mann-Whitney U-test was used to compare differences between groups. PBMCs of healthy subjects (1 × 106/ml, n = 6) were cultured with anti-CD3 (0.25 μg/ml) alone or in combination with IL-12 (20 ng/ml) or IL-21 (50 ng/ml). On day 5, the formation of Tfh cells was recognized by expression of CXCR5 and intracellular production of IL-21. Cells were gated on viable CD4+ lymphocytes. Intensity of CXCR5 expression on survivin+ CD4 cells is shown F. The frequency of CXCR5+ cells within survivin+ and survivin− CD4 subsets stimulated with alpha CD3 + IL-12 G. Intracellular production of IL-21 within the CXCR5+survivin+ and CXCR5+survivin− CD4 cells stimulated with alpha CD3 + IL-12 is shown by histogram H. Frequency of PD-1+ IL-21+ cells is shown by box plots I. The Wilcoxon matched-pairs signed rank test to compare differences. Boxes and lines represent IQR and median, respectively, and error lines indicate min and max values. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/26343374), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Survivin by Flow Cytometry Survivin positive subset of CD44hi CD4 lymphocytes in mouse possess a complete phenotype of Tfh cells Spleen and lymph nodes from collagen II immunized arthritic (CIA) mice were analyzed for expression of survivin and Bcl-6 using flow cytometry A. Cells were gated on memory CD44hiCD4+ lymphocytes. Expression of CXCR5 B. and PD-1 C. was investigated within Bcl-6+ survivin+ and Bcl-6− survivin+ cells. Dots represent individual mice and the horizontal line shows median of the group. Survivin translation in CIA mice was inhibited by shRNA-producing constructs provided as a single intra-peritoneal injection (shSurv16, n = 10, or shSurv13+16, n = 10). Control mice were treated with a non-targeting RNA construct (shNT, n = 9). Survivin expression in spleen was analyzed by flow cytometry 12 days after the injection. Cells were gated on CD4+ lymphocytes. Intensity of survivin expression (MFI) within the groups is shown by a representative histogram and summarized in a box plot D. Survivin expression (MFI) on CD4 lymphocytes correlated to the size of CD44hiCD62L+ population E. Expression of CXCR5 on CD44hiCD4+ lymphocytes in the groups is shown as box plot F. CXCR5+ population correlates with the intensity of survivin G. and with Bcl-6 mRNA H. Boxes and lines represent IQR and median, respectively, and error lines indicate min and max values. The Mann-Whitney U-test was used to compare differences between groups. Correlation analyses were performed using Spearman's test. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/26343374), licensed under a CC-BY license. Not internally tested by R&D Systems.