701760

antibody from Invitrogen Antibodies

Targeting: MET DFNB97, HGFR, RCCP2

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Antibody Data

Product number

701760

Provider

Invitrogen Antibodies

Product name

Phospho-c-Met (Tyr1349) Recombinant Rabbit Monoclonal Antibody (15H7L6)

Antibody type

Monoclonal

Antigen

Synthetic peptide

Reactivity

Human

Host

Rabbit

Isotype

IgG

Antibody clone number

15H7L6

Vial size

100 µg

Concentration

0.5 mg/mL

Storage

Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis was performed on whole cell extracts (50 µg lysate) of MKN 45 (Lane1) and MKN 45 treated with HGF (serum starved/ON, 100 ng/10 min) (Lane 2). The blots were probed with Anti-cMet (pY1349) Recombinant Rabbit Monoclonal Antibody (Product # 701760, 1-2 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:2500 dilution). A 156 kDa band corresponding to cMet (pY1349) was observed according to the treatment. 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® 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).

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunofluorescence was performed on fixed and permeabilized MKN45 cells which were serum starved for 16 hours and treated with 100 ng of HGF/10 min for detection of cMet (pY1349) using Anti-cMet (pY1349) Recombinant Rabbit Monoclonal Antibody (Product # 701760, 1-2 µg/mL) and labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034, 1:2000). Panel a) shows representative cells that were stained for detection and localization of cMet (pY1349) protein (green), Panel b) is stained for nuclei (blue) using SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). Panel c) represents cytoskeletal F-actin staining using Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d) is a composite image of Panels a, b and c clearly demonstrating cytoplasmic localization of cMet (pY1349). Panel e) represents merged image of untreated cells with no signal. Panel f) represents control cells with no primary Antibody to assess background.