Antibody data
- Antibody Data
- Antigen structure
- References [0]
- Comments [0]
- Validations
- Western blot [2]
- Immunocytochemistry [1]
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Validation data
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- Product number
- 720334 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- GAB1 Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Synthetic peptide
- Description
- These Polyclonal antibodies are of rabbit origin developed by immunizing animals with proteins or peptides. The polyclonal antibody is purified by affinity purification from the rabbit sera generated after immunizing the rabbits with a specific type of protein or peptide. The purified antibody is tested for its functionality in various relevant research applications. The antibody is developed for Research Use Only and is non-hazardous or non-infectious in nature.
- Concentration
- 0.5 mg/mL
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-GAB1 Polyclonal Antibody (Product # 720334) and a 90 kDa band corresponding to GAB1 was observed across the panel tested with decrease in expression in U-2 OS cells treated with Anisomycin and in HeLa cells treated with Actinomycin. Whole cell extracts (30 µg lysate) of U-2 OS (Lane 1), U-2 OS (treated with Anisomycin, 50 µm for 8 hr) (Lane 2), HeLa (Lane 3), HeLa ( treated with Actinomycin, 5 µg/mL, 8 hr) (Lane 4), MDA-MB-231 (Lane 5) and SH-SY5Y (Lane 6) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0322BOX). 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 (2 µg/mL) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis was performed on Whole cell extracts (30 µg lysate) of HEK-293 (Lane 1), COS-7 (Lane 2), Hep G2 (Lane 3), SH-SY 5Y (Lane 4) and K-562 (Lane 5). The blots were probed with Anti-Gab1 Rabbit Polyclonal Antibody (Product # 720334, 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 80 kDa band corresponding to Gab1 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® 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
- For immunofluorescence analysis, MCF-7 cells were fixed and permeabilized for detection of endogenous IKK gamma - NEMO using Anti- IKK gamma - NEMO Rabbit Polyclonal Antibody (Product # 720334, 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 IKK gamma - NEMO 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 IKK gamma - NEMO. Panel e) represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.