- 39-2050 - Invitrogen Antibodies VAV1 antibody

Submitted references Involvement of the Rac1-IRSp53-Wave2-Arp2/3 Signaling Pathway in HIV-1 Gag Particle Release in CD4 T Cells.

Thomas A, Mariani-Floderer C, López-Huertas MR, Gros N, Hamard-Péron E, Favard C, Ohlmann T, Alcamí J, Muriaux D
Journal of virology 2015 Aug;89(16):8162-81

Two closely spaced tyrosines regulate NFAT signaling in B cells via Syk association with Vav.

Chen CH, Martin VA, Gorenstein NM, Geahlen RL, Post CB
Molecular and cellular biology 2011 Jul;31(14):2984-96

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on whole cell extracts (30 µg lysate) of Jurkat (Lane 1), THP-1 (Lane 2), T-47D (lane 3) and Raji (Lane 4). The blots were probed with Anti-VAV1 Mouse Monoclonal Antibody (Product # 39-2050, 1:100-1:1000 dilution) and detected by chemiluminescence Goat anti-Mouse IgG (H+L) Secondary Antibody, HRP conjugate (Product # 62-6520, 1:4000 dilution). A 110 kDa band corresponding to VAV1 was observed across cell lines tested. An extra band at ~ 70 kDa was observed in Jurkat and THP-1. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 10 % Bis-Tris gel (Product # NP0301BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane by wet transfer method. 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: Western blot was performed using Anti-VAV1 Monoclonal Antibody (ZV003) (Product # 39-2050) and a 98kDa band corresponding to VAV1 was observed across cell line and tissue tested. Whole cell extracts (30 µg lysate) of Jurkat (Lane 1), MOLT-4 (Lane 2), Raji (Lane 3), THP-1 (Lane 4), HeLa (Lane 5), SH-SY5Y (Lane 6), T-47D (Lane 7) and MCF7 (Lane 8). Tissue extract of Mouse Thymus (Lane 9) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:1000 dilution) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of VAV1 was performed using 70 confluent log phase Jurkat cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with VAV1 Monoclonal Antibody (ZV003) (Product # 39-2050) at 1:100 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Mouse IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32766), (1:2000 dilution), for 45 minutes at room temperature (Panel a: Green). Nuclei (Panel b: Blue) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300 dilution). Panel d represents the merged image showing cytoplasmic localization. Panel e represents no expression in MCF7. Panel f represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

39-2050

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