Isolation of Leishmania Promastigote Flagella

Methods Mol Biol. 2020:2116:485-495. doi: 10.1007/978-1-0716-0294-2_29.

Abstract

Eukaryotic flagella are conserved multifunctional organelles with roles in motility, intercellular interactions, and signal transduction. Leishmania possess a single flagellum at all stages of their life cycle. Flagella of promastigote forms in the fly are long and motile, with a canonical 9 + 2 microtubule axoneme and an extra-axonemal paraflagellar rod (PFR). This protocol describes a simple method for the isolation of Leishmania mexicana promastigote flagella, optimized to yield intact flagella that retain both the cytoskeletal elements (9 + 2 axoneme and PFR) and the surrounding membrane. The isolated flagella and deflagellated cell bodies are suitable for analysis by electron microscopy, protein mass spectrometry, and lipidomics.

Keywords: Cell fractionation; Flagella; Kinetoplastids; Leishmania.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Fractionation / methods*
  • Centrifugation, Density Gradient
  • Cytoskeleton / metabolism
  • Flagella / metabolism*
  • Leishmania mexicana / cytology*
  • Leishmania mexicana / growth & development
  • Life Cycle Stages*
  • Lipidomics
  • Mass Spectrometry
  • Microscopy, Electron
  • Protozoan Proteins / analysis
  • Protozoan Proteins / isolation & purification
  • Protozoan Proteins / metabolism

Substances

  • Protozoan Proteins