Cells were washed twice with cold PBS and lysed in RIPA buffer supplemented with protein and phosphatase inhibitor cocktails. Protein concentration was determined by the Bicinchoninic Acid Assay (Thermo scientific) using BSA as a standard. Equal amounts of protein were mixed with Laemmli buffer 2x warmed at 95°C for 5 min, and loaded on 4-12% gels for SDS-PAGE. After electrophoretic separation, proteins were blotted onto 0.22 mm nitrocellulose (Millipore), blocked with 10% non-fat milk in TBS-Tween, and incubated at 4°C overnight with the following antibodies: anti-succinate dehydrogenase b (goat polyclonal Santa Cruz Biotech sc-34150, 1:500), anti-β-tubulin (mouse monoclonal Sigma-Aldrich, T5201, 1:10000), anti-pyruvate carboxylase (rabbit monoclonal Abcam, ab126707, 1:1000), anti-NDUFS4 (mouse monoclonal, Abcam, Ab87399, 1:1000), anti-MTND6 (mouse monoclonal, Invitrogen, A31857, 1:1000), anti-COX IV (rabbit polyclonal, Abcam, Ab16056, 1:2000), anti-membrane integrity WB cocktail (mouse monoclonal, Abcam, Ab110414, 1:1000). Membranes were then washed and incubated with secondary anti-mouse (926-32212 or 926-32222, LI-COR), anti-rabbit (926-68073, LI-COR) or anti-goat (926-32214, LI-COR) at 1:10000 dilution. The IR scanning was performed using Licor Odyssey scanner (channel, 700 and 800) and acquired using Image Studio 2.0.
Ab110414
Manufactured by Abcam
Ab110414 is a monoclonal antibody that recognizes the human F1-ATPase complex. This antibody can be used for the detection and analysis of the F1-ATPase complex through various immunoassay techniques.
Automatically generated - may contain errors
Lab products found in correlation
Bicinchoninic acid assay, by Thermo Fisher Scientific (2 mentions)
Nitrocellulose, by Merck Group (2 mentions)
Ab16056, by Abcam (2 mentions)
Ab87399, by Abcam (2 mentions)
Ab126707, by Abcam (2 mentions)
Sc 34150, by Merck Group (2 mentions)
T5201, by Abcam (2 mentions)
A31857, by Abcam (2 mentions)
2 protocols using ab110414
1
Mitochondrial protein quantification by Western blot
2
Mitochondrial protein quantification by Western blot
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Cells were washed twice with cold PBS and lysed in RIPA buffer supplemented with protein and phosphatase inhibitor cocktails. Protein concentration was determined by the Bicinchoninic Acid Assay (Thermo scientific) using BSA as a standard. Equal amounts of protein were mixed with Laemmli buffer 2x warmed at 95°C for 5 min, and loaded on 4-12% gels for SDS-PAGE. After electrophoretic separation, proteins were blotted onto 0.22 mm nitrocellulose (Millipore), blocked with 10% non-fat milk in TBS-Tween, and incubated at 4°C overnight with the following antibodies: anti-succinate dehydrogenase b (goat polyclonal Santa Cruz Biotech sc-34150, 1:500), anti-β-tubulin (mouse monoclonal Sigma-Aldrich, T5201, 1:10000), anti-pyruvate carboxylase (rabbit monoclonal Abcam, ab126707, 1:1000), anti-NDUFS4 (mouse monoclonal, Abcam, Ab87399, 1:1000), anti-MTND6 (mouse monoclonal, Invitrogen, A31857, 1:1000), anti-COX IV (rabbit polyclonal, Abcam, Ab16056, 1:2000), anti-membrane integrity WB cocktail (mouse monoclonal, Abcam, Ab110414, 1:1000). Membranes were then washed and incubated with secondary anti-mouse (926-32212 or 926-32222, LI-COR), anti-rabbit (926-68073, LI-COR) or anti-goat (926-32214, LI-COR) at 1:10000 dilution. The IR scanning was performed using Licor Odyssey scanner (channel, 700 and 800) and acquired using Image Studio 2.0.
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