Journal of Clinical and Translational Hepatology

Journal of Clinical and Translational Hepatology

Friday, 07 / 19 / 2019

Articles

An Improved Method for Preparation of Uniform and Functional Mitochondria from Fresh Liver

METHODS ARTICLE

An Improved Method for Preparation of Uniform and Functional Mitochondria from Fresh Liver

Nagham Khouri Farah, Xiaocong Liu*, Catherine H. Wu and George Y. Wu

Department of Medicine, Division of Gastroenterology-Hepatology, University of Connecticut Health Center, Farmington, CT, USA
*Correspondence to: Xiaocong Liu, Department of Medicine, Division of Gastroenterology-Hepatology, University of Connecticut Health Center, Farmington, CT 06030, USA. Tel: +1-860-679-2509, Fax: +860-679-6582, E-mail: This email address is being protected from spambots. You need JavaScript enabled to view it.

Journal of Clinical and Translational Hepatology 2019;7(1):46-50 DOI: 10.14218/JCTH.2018.00064
Received: December 14, 2018 Accepted: February 28, 2019 Published online: March 27, 2019

Abstract
Background and Aims: As the major energy source for mammalian cells, mitochondria have been the subject of numerous studies. However, the isolation and purification of healthy mitochondria, especially from fresh tissue, remains challenging. The most popular methods and kits involve various centrifugation steps which require substantial time and equipment but do not consistently provide pure preparations of functional mitochondria. The aim of this study was to determine whether methods could be devised to improve the purity and yield of functional mitochondria from fresh tissue.

Methods: Fresh mouse liver was homogenized, and cells lysed. Particle size analysis, quantitation of mitochondrial DNA, mitochondrial oxygen consumption, and purity of mitochondria (by electron microscopy) were measured in samples after various purification steps and significant differences determined.

Results: A two-step procedure consisting of centrifugation followed by filtration through 1.2μ and 0.8μ filters resulted in uniform mitochondrial preparations with diameters between 520–540 nm, and approximately 5-times more pure samples. The mitochondria thus obtained had oxygen consumption and sensitivities to mitochondrial inhibitors that were indistinguishable from those purified by centrifugation alone. Electron microscopy confirmed the presence of more uniform and 4–5 times greater concentrations of mitochondria compared to centrifugation alone.

Conclusions: A two-step procedure consisting of sequential centrifugation followed by filtration is a rapid method for the production of highly purified, uniform and functional mitochondria.

Keywords
Mitochondria, Isolation, Filtration, Purified, Uniform, Functional

 

 

 

Journal of Clinical and Translational Hepatology 2019 vol. 7, 46-50  [ Html ] [ PDF Full-text ]

 

© The Authors 2018. This article is published under the terms of the Creative Commons Attribution-Noncommercial License (CC BY-NC 4.0), which permits use, distribution, and reproduction in any medium, provided the original work is properly cited, the use is non commercial and is otherwise in compliance with the license.

 

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