A Scalable Purification Method for Mitochondria from Trypanosoma brucei

Methods Mol Biol. 2020;2116:611-626. doi: 10.1007/978-1-0716-0294-2_36.

Abstract

Due to its unique biology the mitochondrion of Trypanosoma brucei has attracted a lot of interest since many decades, making it arguably the best studied mitochondrion outside yeast and mammals. Here we describe a method allowing purification of mitochondria from procyclic trypanosomes that yields highly enriched and functional organelles. The method is based on isotonic lysis of cells by nitrogen cavitation, DNase I digestion, differential centrifugation and Nycodenz gradient centrifugation. The method is scalable and can be adapted to culture volumes a small as 100 mL or as large as 24 L.

Keywords: Mitochondria; Nitrogen cavitation; Subcellular fractionation; Trypanosoma brucei; Trypanosomes.

MeSH terms

  • Cell Fractionation / instrumentation
  • Cell Fractionation / methods*
  • Centrifugation, Density Gradient / instrumentation
  • Centrifugation, Density Gradient / methods
  • Mitochondria*
  • Trypanosoma brucei brucei / cytology*