MA5-15227

antibody from Invitrogen Antibodies

Targeting: MAPK3 ERK1, p44erk1, p44mapk, PRKM3

Western blot (Supportive data in Antibodypedia)

Antibody Data

Product number

MA5-15227

Provider

Invitrogen Antibodies

Product name

ERK1/ERK2 Monoclonal Antibody (E.657.8)

Antibody type

Monoclonal

Antigen

Synthetic peptide

Description

It is not recommended to aliquot this antibody. This antibody is not cross-reactive with either JNK/SAPK or p38 MAP kinase.

Reactivity

Human, Mouse, Rat, Bovine, Hamster, Porcine, Zebrafish

Host

Mouse

Isotype

IgG

Antibody clone number

E.657.8

Vial size

100 µL

Concentration

75 µg/mL

Storage

-20°C

Western blot

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.