Mitochondrial Isolation: A Technique to Isolate Mitochondria from Human Ovarian Cancer Tissue Sample
Published: April 30, 2023
Abstract
Source: Zhan, X. et al. Preparation of Mitochondria from Ovarian Cancer Tissues and Control Ovarian Tissues for Quantitative Proteomics Analysis. J. Vis. Exp. (2019)
This video describes the procedure for isolating mitochondria from human ovarian cancer tissues using differential-speed centrifugation in combination with density gradient centrifugation. The isolated mitochondria can be used for proteomic analysis of human ovarian cancer mitochondrial proteome.
Protocol
All procedures involving human participants have been performed in compliance with the institutional, national, and international guidelines for human welfare and have been reviewed by the local institutional review board. 1. Preparation of mitochondria from human ovarian cancer tissues Ovarian tissue samples including ovarian cancer tissues (n = 7) were used for this protocol. Prepare 250 mL of the mitochondrial isolation buffer by mixi…
Declarações
The authors have nothing to disclose.
Materials
| Diamine tetraacetic acid (EDTA) | Sigma | 798681-100G | Anhydrous, free-flowing, Redi-Dri, ≥98% |
| DTT | Sigma | 10197777001 | 1,4-Dithiothreito |
| Ethylen glycol bis(2-aminoethyl ether)tetraacetic acid (EGTA) | Sigma | E0396-10G | BioXtra, ≥97 .0% |
| Homogenizer | SilentShake | HYQ-3110 | |
| Low-temperature super-speed centrifuger | Eppendorf | 5424R | |
| Centrifuge | XiangYi | TDZ4–WS | |
| Mannitol | Macklin | M813424-100G | Mannitol is a polyol (polyhydric alcohol) produced from hydrogenation from fructose that functions as a sweetener, humectant, and bulking agent. It has low hygroscopicity and poor oil solvency. |
| Phenylmethanesulfonyl fluoride (PMSF) protease inhibitor | Solarbio | P0100-1ML | PMSF is a protease inhibitor that reacts with serine residues to inhibit trypsin, chymotrypsin, thrombin, and papain. |
| Potassium chloride | Macklin | P816354-25G | Potassium chloride, KCI, also known as potassium muriate and sylvite, is a colorless crystalline solid with a salty taste that melts at 776°C (1420 OF). It is soluble in water, but insoluble in alcohol. Potassium chloride is used in fertilizers, pharmaceuticals, photography, and as a salt substitute |
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Mitochondrial Isolation: A Technique to Isolate Mitochondria from Human Ovarian Cancer Tissue Sample. J. Vis. Exp. (Pending Publication), e20370, doi: (2023).