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Western blotAntibody data
- Antibody Data
- Antigen structure
- References [3]
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- Validations
- Western blot [3]
- Immunocytochemistry [1]
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- Product number
- MA5-12800 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- GNAI1 Monoclonal Antibody (R4.5)
- Antibody type
- Monoclonal
- Antigen
- Purifed from natural sources
- Description
- MA5-12800 targets Galphai1 G-Protein in WB applications and shows reactivity with Bovine, Guinea Pig, Human, mouse, and Rat samples.
- Antibody clone number
- R4.5
- Concentration
- 0.2 mg/mL
Submitted references Identification of residues crucial for the interaction between human neuroglobin and the α-subunit of heterotrimeric Gi protein.
Human neuroglobin functions as an oxidative stress-responsive sensor for neuroprotection.
Molecular basis of guanine nucleotide dissociation inhibitor activity of human neuroglobin by chemical cross-linking and mass spectrometry.
Takahashi N, Wakasugi K
Scientific reports 2016 Apr 25;6:24948
Scientific reports 2016 Apr 25;6:24948
Human neuroglobin functions as an oxidative stress-responsive sensor for neuroprotection.
Watanabe S, Takahashi N, Uchida H, Wakasugi K
The Journal of biological chemistry 2012 Aug 31;287(36):30128-38
The Journal of biological chemistry 2012 Aug 31;287(36):30128-38
Molecular basis of guanine nucleotide dissociation inhibitor activity of human neuroglobin by chemical cross-linking and mass spectrometry.
Kitatsuji C, Kurogochi M, Nishimura S, Ishimori K, Wakasugi K
Journal of molecular biology 2007 Apr 20;368(1):150-60
Journal of molecular biology 2007 Apr 20;368(1):150-60
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot of Galphai1 G-Protein using Galphai1 G-Protein Monoclonal Antibody (Product # MA5-12800) on IMR-5 Cells.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis was performed on whole cell extracts (30 µg lysate) of IMR-32 (Lane 1), A549 (Lane 2), SH-SY5Y (Lane 3), RSC96 (Lane 4) and tissue extract (30 µg lysate) of Rat Brain (Lane 5).The blots were probed with GNAI1 (Galphai1) Mouse Monoclonal Antibody (Product # MA5-12800, 2 µg/mL) 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).A 40 kDa band corresponding to GNAI1 (Galphai1) was observed across the cell lines and tissue tested. Known quantity of protein samples were electrophoresed using Novex®NuPAGE®4-12 % Bis-Tris gel (Product # NP0322BOX), 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).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Knockdown of GNAI1 was achieved by transfecting A549 cells with GNAI1 specific siRNAs (Silencer® select Product # s5872, s5873). Western blot analysis (Fig a) was performed using whole cell extracts from the GNAI1 knockdown cells (lane 2) and non-specific scrambled siRNA transfected cells (lane 1). The blots were probed with Anti-GNAI1 Monoclonal Antibody (Product # MA5-12800, 1:4000) and Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.25 µg/mL, 1:4000 dilution). Densitometric analysis of this western blot is shown in histogram (Fig b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to GNAI1.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of Galphai1 (GNAI1) was performed using 70% confluent log phase RSC-96 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with GNAI1 (R4.5) Mouse Monoclonal Antibody (Product # MA5-12800) at 2 µg/mL in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Mouse IgG (H+L) Superclonal Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing membranous localization. Panel e shows the no primary antibody control. The images were captured at 60X magnification.
