The methods used for preparation of cell lysates from in vitro and in vivo samples and for immunoblot analyses have been described previously15 (link). The antibodies used include Drp1 (BD Transduction, 611112), phospho-Drp1 (Ser616) (Cell Signaling, 4494S), phospho-Drp1 (Ser637) (Cell Signaling, 4867S), LC3 (BML, M186-3), p62 (ORIGENE, TA307334), Cox I (abcam, ab110413), Beclin 1 (Cell signaling, 3738S), PGC-1α (Santa Cruz, sc-13067), Cox IV (Cell Signaling, 4844S), Mst1 (BD Transduction, 611052), phospho-Mst1 (Cell Signaling, 3681S) and GAPDH (Cell Signaling, 2118S).
Cox 1
Manufactured by Abcam
COX I is a protein that is a subunit of the enzyme complex cytochrome c oxidase, which is involved in the electron transport chain and plays a crucial role in cellular respiration. This product is a research-use-only reagent, and its core function is to facilitate the study of cytochrome c oxidase and its role in cellular processes.
Automatically generated - may contain errors
Lab products found in correlation
Coxii, by Abcam (2 mentions)
Cox 4, by Abcam (2 mentions)
Cox 4, by Cell Signaling Technology (1 mentions)
Beclin 1, by Cell Signaling Technology (1 mentions)
Pgc 1α, by Santa Cruz Biotechnology (1 mentions)
Phospho drp1 ser 637, by Cell Signaling Technology (1 mentions)
Phospho drp1 ser616, by Cell Signaling Technology (1 mentions)
Phospho mst1, by Cell Signaling Technology (1 mentions)
Gapdh, by Cell Signaling Technology (1 mentions)
Bcl xl, by Abcam (1 mentions)
Cd133, by Aviva Systems Biology (1 mentions)
P drp1, by Abcam (1 mentions)
Gapdh, by Abcam (1 mentions)
Bcl2 associated x (bax), by Abcam (1 mentions)
Caspase 3, by Abcam (1 mentions)
Anti sdha, by Abcam (1 mentions)
Micu1, by Cell Signaling Technology (1 mentions)
3 protocols using cox 1
1
Cell Lysate Preparation and Immunoblot Analysis
2
Mitochondrial Dynamics and EV Profiling
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Mitochondria were isolated from PK1 cells using a mitochondria isolation kit (Thermos Fisher, USA), following vendor’s instructions, as we have shown37 . Protein concentration was measured with Bradford Protein Assay. Specific antibodies against Drp1 (1:1000; Abcam), Mfn2 (1:1000; Abcam) and OPA1 (1:500; Abcam) were used with blotting protocols. COXIV (1:5000; Abcam) was used as loading controls. Cell surface, EV or kidney protein expression was studied in homogenate. Specific antibodies against CD9 (1:5,000; Abcam), CD81 (1:1,000; Abcam), CD29(1:1000, Abcam), CD24(1:500, Abcam), CD133(1:500, Aviva Systems Biology), COX I (1:1000, Abcam), COX II (1:1000,Abcam), Caspase3 (1:1000, Abcam), Bax(1:1000,Abcam), Bcl-XL (1:1000, Abcam), COX IV, Drp1, p-Drp1(1:1000, Abcam) antibodies were used with blotting protocols, and GAPDH (1:1000, Abcam) as loading controls.The density of each band was analyzed by Image-Pro Plus 6.0 software.
3
Assessing Mitochondrial OXPHOS Complex Activity
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A spectrophotometric assay of whole-cell extracts from control and shRNAtreated cells was used to measure the enzymatic activity of COX In fibroblasts as described previously (Antonicka et al., 2003) .
Native, Denaturing, and Two-Dimensional PAGE Blue native PAGE (BN-PAGE) was used to separate individual OXPHOS complexes. Mitoplasts, prepared from fibroblasts by treatment with 0.8 mg of digitonin per milligram of protein, were solubilized with 1% lauryl maltoside. Samples (10-20 mg) were run on 6%-15% polyacrylamide gradient gels as described in detail elsewhere (Leary, 2012) . The following antibodies were used in this study: anti-CCDC90A (LS-Bioscience and AVIVA), anti-SDHA, a-actin, COX I, COX II, COX IV, PDH (Abcam), MCU (1/500 dilution), P-PDH (Sigma), and MICU1 (1/500 dilution; Cell Signaling).
Native, Denaturing, and Two-Dimensional PAGE Blue native PAGE (BN-PAGE) was used to separate individual OXPHOS complexes. Mitoplasts, prepared from fibroblasts by treatment with 0.8 mg of digitonin per milligram of protein, were solubilized with 1% lauryl maltoside. Samples (10-20 mg) were run on 6%-15% polyacrylamide gradient gels as described in detail elsewhere (Leary, 2012) . The following antibodies were used in this study: anti-CCDC90A (LS-Bioscience and AVIVA), anti-SDHA, a-actin, COX I, COX II, COX IV, PDH (Abcam), MCU (1/500 dilution), P-PDH (Sigma), and MICU1 (1/500 dilution; Cell Signaling).
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