The primary antibodies used were: mouse monoclonal anti-Rab2 (Abcam), rabbit polyclonal anti-Sar1 (Abcam), mouse monoclonal anti-GAPDH (Cell Signaling Technology), mouse monoclonal anti-His(Sigma), mouse monoclonal anti-Flag (Sigma), mouse monoclonal anti-Myc (Cell Signaling Technology), mouse monoclonal anti-HA (Cell Signaling Technology), mouse monoclonal anti-GroEL(Santa Cruz) and mouse monoclonal anti-Omp1 (Santa Cruz). The secondary antibodies used for Western blotting were: Horse Radish Peroxidase (HRP)-conjugated Goat Anti-Mouse IgG (Santa Cruz) and HRP-conjugated Goat Anti-Rabbit IgG (Cell Signaling) antibodies. Normal rabbit IgG was purchased from Santa Cruz. The secondary antibodies used for immunofluorescence were: goat anti-rabbit IgG (H + L) secondary antibody, Alexa Fluor® 488 conjugate (Invitrogen), and donkey anti-rabbit IgG secondary antibody and Alexa Fluor® 594 conjugate (Invitrogen).
Mouse monoclonal anti gapdh
Manufactured by Cell Signaling Technology
Sourced in United States
Mouse monoclonal anti-GAPDH is a primary antibody that specifically binds to the GAPDH (Glyceraldehyde 3-phosphate dehydrogenase) protein. GAPDH is a highly conserved enzyme involved in the glycolytic pathway.
Automatically generated - may contain errors
Lab products found in correlation
Hrp conjugated goat anti rabbit igg, by Cell Signaling Technology (1 mentions)
Alexa fluor 488 conjugate, by Thermo Fisher Scientific (1 mentions)
Mouse monoclonal anti his, by Merck Group (1 mentions)
Mouse monoclonal anti myc, by Cell Signaling Technology (1 mentions)
Mouse monoclonal anti ha, by Cell Signaling Technology (1 mentions)
Horseradish peroxidase hrp conjugated goat anti mouse igg, by Santa Cruz Biotechnology (1 mentions)
Alexa fluor 594 conjugate, by Thermo Fisher Scientific (1 mentions)
Mouse monoclonal anti flag, by Merck Group (1 mentions)
Radioimmunoprecipitation assay buffer, by Boston BioProducts (1 mentions)
Rabbit polyclonal anti akt, by Cell Signaling Technology (1 mentions)
Nupage lds sample buffer 4, by Thermo Fisher Scientific (1 mentions)
Rabbit monoclonal anti p akt, by Cell Signaling Technology (1 mentions)
Lipod293 in vitro dna transfection reagent, by SignaGen (1 mentions)
Puromycin, by Merck Group (1 mentions)
Actinomycin d, by Merck Group (1 mentions)
Hygromycin b, by Merck Group (1 mentions)
Chemiluminescence kit, by Merck Group (1 mentions)
Pvdf membrane, by Merck Group (1 mentions)
Ripa buffer, by Beyotime (1 mentions)
Bca protein assay kit, by Beyotime (1 mentions)
5 protocols using mouse monoclonal anti gapdh
1
Comprehensive Immunoblotting and Immunofluorescence Protocols
2
Comprehensive Antibody and Reagent List
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Specific reagents used in this study were actinomycin D, G418, hygromycin B, puromycin, sodium butyrate, TriPure Isolation Reagent solution, and valproic acid from Millipore Sigma, doxycycline and NuPAGE™ LDS Sample Buffer (4×) from Thermo Fisher Scientific, radioimmunoprecipitation assay buffer (RIPA) from Boston Bioproducts (Ashland, MA), and LipoD293™ In Vitro DNA Transfection Reagent from SignaGen Laboratories (Frederick, MD). Following antibodies were used in this report: affinity-purified rabbit polyclonal anti-ORF57 N-terminal (in house), mouse monoclonal anti-ORF57 N-terminal, and affinity-purified rabbit polyclonal anti-ORF57 C-terminal antibodies were from Rockland Immunochemicals. Rabbit polyclonal anti-AKT, rabbit monoclonal anti-p-AKT, rabbit monoclonal anti-c-FOS (9F6), mouse monoclonal anti-GAPDH, and rabbit polyclonal anti-c-JUN antibodies were purchased from Cell Signaling Technology. Rabbit polyclonal anti-RTA was a gift of Dr. Yoshi Izumiya (UC Davis). Mouse monoclonal anti-myc (9E10) was a gift of Dr. Xuefeng Liu (the Ohio State University). Mouse polyclonal anti-AEN was from Millipore Sigma, rabbit polyclonal anti-RGS2 from Abcam, rabbit polyclonal anti-ZFC3H1(MTR4) antibody from Novus biologicals, and rabbit IgG isotype, mouse IgG isotype were from Thermo Fisher Scientific.
3
Western Blot Analysis of γ-H2AX
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Total proteins were extracted from cultured cells using RIPA buffer (Beyotime) containing protease inhibitors cocktail (Roche, Switzerland). Protein concentration was quantified using the BCA protein assay kit (Beyotime). Equal amounts of protein were subjected to 12% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and transferred onto polyvinylidene difluoride (PVDF) membrane (Millipore, Billerica, MA). After being blocked with 5% nonfat milk in TBST for 1 h at room temperature, the membranes were incubated with primary antibodies, including mouse monoclonal anti-γ-H2AX (No. 2577; 1: 1000 dilution, Cell Signaling Technology, Beverly, MA) and mouse monoclonal anti-GAPDH (No. 97166; 1: 2000 dilution, Cell Signaling Technology) at 4°C overnight, and then by probed with horseradish peroxidase (HRP)-conjugated goat anti-rabbit IgG (sc-2379; 1: 1000 dilution, Santa Cruz Biotechnology, Santa Cruz, CA) for 2 h. Reactive protein expression was visualized and detected using a chemiluminescence kit (Millipore).
4
Western Blot Analysis of Apoptosis Regulators
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INS-1 cells and mouse pancreases were lysed in radioimmunoprecipitation assay (RIPA) buffer (Pierce Biotechnology Corporation, Rockford, IL, USA), and quantified for total protein using a Micro BCA Protein Assay Kit (Pierce Technology). Total proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and then transferred to a polyvinylidene fluoride membrane (Bio-Rad Laboratories, Hercules, CA, USA). The membrane was then blocked with 5% skimmed milk. The membrane was incubated overnight at 4 °C with one of the following primary antibodies: mouse monoclonal anti-TRAIL, goat polyclonal anti-DR5, goat polyclonal anti-DcR1, mouse monoclonal anti-Bax, rabbit polyclonal anti-Bcl-2, mouse monoclonal anti-NF-κB, goat polyclonal anti-α-tubulin, mouse monoclonal anti-β-actin (all from Santa Cruz Biotechnology, Dallas, TX, USA), or mouse monoclonal anti-GAPDH (Cell Signaling Technology, Danvers, MA, USA). The membranes were then washed and incubated with horseradish peroxidase (HRP)-conjugated secondary antibody (Santa Cruz Biotechnology). Protein bands were detected by enhanced chemiluminescence (Pierce Biotechnology). The band intensities of the proteins were analyzed using ImageJ densitometry software (version 1.43; National Institute of Health, Bethesda, MD, USA).
5
Western Blot Analysis of Cellular Proteins
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Whole-cell lysates were obtained using the ProteoJET Mammalian Cell Lysis Reagent (Thermo Scientific, Rockford, IL, USA). Proteins (50 μg) were separated on 10% SDS-PAGE gels and transferred to nitrocellulose membranes. The membranes were blocked with 5% non-fat dried milk in TBST (50 mM Tris (pH 7.5), 200 mM NaCl, 0.05% Tween 20) at room temperature for 1 h. The membranes were then incubated with the primary antibody in the blocking solution for 1 h at room temperature, washed three times with TBST for 15 min, incubated with the HRP-conjugated secondary antibody at room temperature for 1 h and then washed three times with TBST. Antigen-antibody complexes were detected using the SuperSignal detection reagents (Thermo Scientific, Rockford, IL, USA). The following antibodies were used: rabbit monoclonal anti-p53, rabbit monoclonal anti-AGO2, rabbit monoclonal anti-GADD45A, mouse monoclonal anti-GAPDH (Cell Signalling Technology, MA, USA) and horseradish peroxidase-conjugated goat-anti-rabbit and goat-anti-mouse secondary antibodies (Santa Cruz. CA, USA).
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