Anti-CD146 MCAM Rabbit Monoclonal Antibody

Western blot analysis of CD146 using anti-CD146 antibody (M01683-1).
Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 30 ug of sample under reducing conditions.
Lane 1: human U2OS whole cell lysates,
Lane 2: human K562 whole cell lysates,
Lane 3: human Hela whole cell lysates,
Lane 4: human A549 whole cell lysates,
Lane 5: rat lung tissue lysates,
Lane 6: rat spleen tissue lysates,
Lane 7: mouse lung tissue lysates.
After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% non-fat milk/TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-CD146 antigen affinity purified monoclonal antibody (Catalog # M01683-1) at 1:500 overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:500 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for CD146 at approximately 120 kDa. The expected band size for CD146 is at 72 kDa.

IHC analysis of CD146 using anti-CD146 antibody (M01683-1).
CD146 was detected in a paraffin-embedded section of human melanoma tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1:50 rabbit anti-CD146 Antibody (M01683-1) overnight at 4°C. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using HRP Conjugated Rabbit IgG Super Vision Assay Kit (Catalog # SV0002) with DAB as the chromogen.

IHC analysis of CD146 using anti-CD146 antibody (M01683-1).
CD146 was detected in a paraffin-embedded section of human placenta tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1:50 rabbit anti-CD146 Antibody (M01683-1) overnight at 4°C. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using HRP Conjugated Rabbit IgG Super Vision Assay Kit (Catalog # SV0002) with DAB as the chromogen.

Immunofluorescent analysis of A375 cells, using CD146 Antibody.

IF analysis of CD146 using anti-CD146 antibody (M01683-1). CD146 was detected in a paraffin-embedded section of zebrafish brain tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 5% BSA. The tissue section was then incubated with rabbit anti-CD146 Antibody (M01683-1) at a dilution of 1:50 overnight at 4°C. DyLight®488 Conjugated Goat Anti-Rabbit IgG (BA1127) was used as secondary antibody at 1:500 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.