Human/Mouse RUNX3/CBFA3 Antibody

Detection of RUNX3/CBFA3 in Human PBMC by Flow Cytometry. Human PBMC unstimulated (light orange filled histogram) or treated with 50ng/mL PMA (dark orange filled histogram) were stained with Mouse Anti-Human/Mouse RUNX3/CBFA3 Monoclonal Antibody (Catalog # MAB3765) or isotype control antibody (Catalog # MAB003, open histogram), followed by Phycoerythrin-conjugated Anti-Mouse IgG F(ab')₂Secondary Antibody (Catalog # F0102B). To facilitate intracellular staining, cells were fixed with paraformaldehyde and permeabilized with saponin.

RUNX3/CBFA3 in Human PBMCs. RUNX3/CBFA3 was detected in immersion fixed human peripheral blood mononuclear cells (PBMCs) using Mouse Anti-Human/Mouse RUNX3/CBFA3 Monoclonal Antibody (Catalog # MAB3765) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Mouse IgG Secondary Antibody (red; Catalog # NL007) and counterstained with DAPI (blue). Specific staining was localized to nuclei. View our protocol for Fluorescent ICC Staining of Non-adherent Cells.

Detection of Human and Mouse RUNX3/CBFA3 by Western Blot. Western blot shows lysates of Jurkat human acute T cell leukemia cell line, Daudi human Burkitt's lymphoma cell line, and DA3 mouse myeloma cell line. Gels were loaded with 30 µg of whole cell lysate (WCL), 20 µg of cytoplasmic (Cyto), and 10 µg of nuclear extracts (Nuc). PVDF Membrane was probed with 1 µg/mL of Mouse Anti-Human/Mouse RUNX3/CBFA3 Monoclonal Antibody (Catalog # MAB3765) followed by HRP-conjugated Anti-Mouse IgG Secondary Antibody (Catalog # HAF007). Specific bands were detected for RUNX3/CBFA3 at approximately 48 and 52 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Detection of Human RUNX3/CBFA3 by Simple Western^TM. Simple Western lane view shows lysates of Jurkat human acute T cell leukemia cell line and Daudi human Burkitt's lymphoma cell line, loaded at 0.5 mg/mL. A specific band was detected for RUNX3/CBFA3 at approximately 54 kDa (as indicated) using 10 µg/mL of Mouse Anti-Human/Mouse RUNX3/CBFA3 Monoclonal Antibody (Catalog # MAB3765). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.

Detection of RUNX3/CBFA3 by Flow Cytometry RA and RAR alpha reciprocally regulate the expression of the positive and negative LC-regulating transcription factors, Runx3 and C/EBP beta. a Impact of RA and RAR alpha deficiency on Runx3 expression at mRNA level. b Impact of RA and RAR alpha deficiency on Runx3 expression at protein level. c Effect of enforced Runx3 expression on BM-derived LC differentiation in the presence and absence of RA. The data shown are gated for transduced Thy1.1+CD11c+ cells. d Impact of RA and RAR alpha deficiency on expression of Cebpb mRNA. e Impact of RA and RAR alpha deficiency on expression of C/EBP beta protein. f Effect of dnC/EBP beta on BM-derived LC differentiation in the presence and absence of RA. BM cells from WT or ∆RaraCD11c mice were cultured with GM-CSF and TGF beta 1 for 5 days (3 days following retroviral transduction) in the presence of At-RA (10 nM except in panels c and f where 0.1 nM was used) in a medium containing charcoal-treated FBS. Representative and combined data (n = 3–7) from at least 3 experiments are shown. Significant differences from controls by one-way ANOVA with Bonferroni adjustments (p < 0.05)* or between indicated groups by two-way ANOVA with Tukey adjustments (p < 0.05)**. Error bars are defined as s.e.m Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/30254197), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of RUNX3/CBFA3 by Flow Cytometry RA regulates CEBPb and RUNX3 expression and human langerin+ cell differentiation from blood monocytes. a Human LC differentiation from blood monocytes in regular vs. charcoal FBS. b At-RA promotes C/EBP beta + non-LC differentiation. c RA and RAR alpha antagonists reciprocally regulate human LC differentiation. d Effects of At-RA and BMS493 on the expression of human CEBPb and RUNX3. Human blood CD14+ monocytes were cultured in GM-CSF and TGF-beta 1 for 5–7 days in media containing charcoal FBS except in panel A where regular FBS was also used. At-RA was used at 1 nM, and BMS493 and Ro4153 were used at 500 nM. Representative and combined data (n = 5–7) from at least 5 experiments are shown. *Significant differences from control by Mann–Whitney U test (p < 0.05, unpaired, 2-sided). Error bars are defined as s.e.m. Langerhans cells (LC) and langerin-expressing conventional dendritic cells are made from distinct progenitors and enriched in the distinct microenvironments of the skin. Here the authors show that these immune cells are regulated by RAR alpha via simultaneous induction of LC-promoting Runx3 and repression of LC-inhibiting C/EBP beta Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/30254197), licensed under a CC-BY license. Not internally tested by R&D Systems.