Mouse MCAM/CD146 Antibody

Detection of MCAM/CD146 in NK1.1+ Mouse Splenocytes by Flow Cytometry.

Mouse NK1.1+ splenocytes were stained with Rat Anti-Mouse MCAM/CD146 Monoclonal Antibody (Catalog # MAB7718, filled histogram) or isotype control antibody (Catalog # MAB006, open histogram), followed by Allophycocyanin-conjugated Anti-Rat IgG Secondary Antibody (Catalog # F0113).

MCAM/CD146 in bEnd.3 Mouse Cell Line.

MCAM/CD146 was detected in immersion fixed bEnd.3 mouse endothelioma cell line using Rat Anti-Mouse MCAM/CD146 Monoclonal Antibody (Catalog # MAB7718) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the Northern-Lights™ 557-conjugated Anti-Rat IgG Secondary Antibody (red; Catalog # NL013) and counterstained with DAPI (blue). Specific staining was localized to cell surfaces and cytoplasm. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

Detection of Mouse MCAM/CD146 by Immunocytochemistry/Immunofluorescence

MCAM is required to establish cell autonomous polarity. (A) In elongating myotubes (10T1/2 cells treated with testosterone for 7 days) VANGL2 is localized asymmetrically at the tip of the cell. (B) The VANGL2 enriched tip of the cell is marked by MSN. (C) In MCAM knockout C164 cells myotube elongation fails, MSN labels the whole plasma membrane and VANGL2 is spread across the cytoplasm. (D) Highly polarized localization of MCAM and SCRIB at the distal end of growing wild-type myotube. Separate channels of the boxed area are shown on the right. (E) In MCAM knockout cells SCRIB levels remain low and it is spread evenly in the cell. (F) In wild-type myotubes PAR3 remains cytoplasmic, whereas (G) in MCAM knockout C164 cells it can be detected at the cell cortex. (H) RT-qPCR demonstrates reduced expression of Scrib in MCAM mutant cell lines. Cells were treated for 7 days with BMP2 or testosterone (n=3; **P0.01; ***P0.001; two-tailed t-test; mean±s.e.m.). (I) Deletion of MCAM endocytosis motif leads to similar polarity defects as complete MCAM elimination. VANGL2 is evenly spread in U125 cells and PAR3 accumulates in cell cortex. (J) In chondrogenic differentiation VANGL2 was observed asymmetrically in limited number of cells. In MCAM mutant cell lines (C149, C164, U125) VANGL2 accumulated around the nucleus. (K) Initiation of myogenic (4-day culture with testosterone) and chondrogenic differentiation (4-day culture with BMP2) led to downregulation of ERK1/2 phosphorylation (p-ERK1/2). Instead in MCAM mutant cell lines ERK1/2 phosphorylation increased. Scale bars: 25 µm. Image collected and cropped by CiteAb from the following publication (https://journals.biologists.com/bio/article/doi/10.1242/bio.027771/256759/MCAM-contributes-to-the-establishment-of-cell), licensed under a CC-BY license. Not internally tested by R&D Systems.