MA5-15227

antibody from Invitrogen Antibodies

Targeting: MAPK3 ERK1, p44erk1, p44mapk, PRKM3

Antibody data

Submit

Validation data

MAPK3 protein structure - MA5-15227 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on membrane enriched extracts (30 µg lysate) of HeLa (Lane 1), H1975 (Lane 2), HCT 116 (Lane 3), HT-29 (Lane 4), A-431 (Lane 5), A549 (Lane 6), MDA-MB-231 (Lane 7), MCF7 (Lane 8) and U-87 MG (Lane 9). The blot was probed with Mouse Anti-ERK1/ERK2 Monoclonal Antibody (Product # MA5-15227, 1:2000) and detected by chemiluminescence using Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.25 µg/mL, 1:4000 dilution). 42, 44 kDa band corresponding to ERK1/ERK2 was observed across the cell lines tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0321BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5 % skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockdown of ERK1/ERK2 was achieved by transfecting MCF7 cells with ERK1/ERK2 specific validated siRNAs (Silencer® select Product # s11140, s11137 and s11138). Western blot analysis (Fig a) was performed using whole cell lysates from the ERK1/ERK2 knock down cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blots were probed with Anti-ERK1/ERK2 Mouse Monoclonal Antibody (Product # MA5-15227, 1:500) and Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.4 µg/mL, 1:4000 dilution). Densitometric analysis of this western blot is shown in histogram (Fig b). Loss of signal upon siRNA mediated knock down confirms that antibody is specific to ERK1/ERK2.