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
- 700727 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- GATA1 Recombinant Rabbit Monoclonal Antibody (4H15L5)
- Antibody type
- Monoclonal
- Antigen
- Recombinant full-length protein
- Reactivity
- Human, Mouse
- Host
- Rabbit
- Isotype
- IgG
- Antibody clone number
- 4H15L5
- 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.
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-GATA1 Recombinant Rabbit Monoclonal Antibody (4H15L5) (Product # 700727) and a 50kDa band corresponding to Erythroid transcription factor was observed across in K-562 and HEL 92.1.7 cell lines as opposed to all other models tested. Nuclear enriched extracts (30 µg lysate) of HEL 92.1.7 (Lane 1), K-562 (Lane 2), Jurkat (Lane 3), Raji (Lane 4), A-431 (Lane 5), SK-O-V3 (Lane 6), IMR-32 (Lane 7), Mouse Thymus (Lane 8), Mouse Spleen (Lane 9), HeLa (Lane 10) 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 # A27036,1:20,000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using SuperSignal™ West Pico PLUS Chemiluminescent Substrate (Product # 34580).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of GATA1 was performed by loading 20 µg of K562 cell lysate using Novex®NuPAGE® 4-12% Bis-Tris gel (Product # NP0321BOX), XCell SureLock Electrophoresis System (Product # EI0002), Novex® Sharp Pre-Stained Protein Standard (Product # LC5800), and iBlot® Dry Blotting System (Product # IB21001). Proteins were transferred to a nitrocellulose membrane and blocked with 5% skim milk for 1 hour at room temperature. GATA1 was detected at ~ 50 kDa using GATA1 Recombinant Rabbit Monoclonal Antibody (Product # 700727) at 1-3 µg/mL in 2.5% skim milk at 4°C overnight on a rocking platform. Goat anti-Rabbit IgG - HRP Secondary Antibody (Product # G-21234) at 1:5000 dilution was used and chemiluminescent detection was performed using Pierce™ ECL Western blotting Substrate (Product # 32106).
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of Erythroid transcription factor was performed using log phase K-562 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with GATA1 Recombinant Rabbit Monoclonal Antibody (4H15L5) (Product # 700727) at 1:100 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 594 (Product # A32754), (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 withRhodamine Phalloidin (Product # R415, 1:300 dilution). Panel d represents the merged image showing nuclear localization. Panel e shows Jurkat cells with no expression of GATA1. Panel f represents control cells with no primary antibody to assess background. The images were captured at 60X magnification in EVOS™ M7000 Imaging System (Product # AMF7000) and externally deconvoluted (D.Sage et al. / Methods 115 (2017) 28-41).