Establishing a liquid-phase IEF in combination with 2-DE for the analysis of Leishmania proteins

Reynolds K.B. Brobey, Lynn Soong

Research output: Contribution to journalArticle

23 Scopus citations

Abstract

The recent completion of genome sequencing projects for Leishmania major and near completion for two other species, L. infantum and L. braziliensis, has provided the needed genomic information for investigating the proteomes of Leishmania parasites. However, the design of effective 2-DE-based proteome mapping for complex protozoan parasites like Leishmania has proven to be severely compromised due to extensive overcrowding of spots especially in the acidic regions, coupled to a relatively low representation of basic proteins. In the present study, we optimized a liquid-phase IEF in combination with 2-DE for L. amazonensis promastigote as a way of reducing protein complexity and enhancing representation for low-abundance proteins on gels. Of 20 pH-based fractions eluted from Rotofor™ cells, 5 representative fractions selected from acidic, basic or neutral regions of the proteome and with adequate protein concentration were further analyzed by 2-DE using medium-range IPG strips. On this basis, we were able to generate high-resolution 2-DE maps encompassing both the acidic and basic ends of the proteome with enhanced spot representation.

Original languageEnglish (US)
Pages (from-to)116-120
Number of pages5
JournalProteomics
Volume7
Issue number1
DOIs
StatePublished - Jan 1 2007

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Keywords

  • Leishmania amazonensis
  • Liquid-phase IEF
  • Parasite
  • Proteome
  • Rotofor cell

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

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