A mass spectrometry-based workflow for the proteomic analysis of in vitro cultured cell subsets isolated by means of laser capture microdissection

Research output: Contribution to journalArticle

Abstract

This paper describes a microproteomic workflow that is useful for simultaneously identifying and quantifying proteins from a minimal number of morphotypically heterogeneous cultured adherent cells. The analytical strategy makes use of laser capture microdissection, an effective means of harvesting pure cell populations, and label-free mass spectrometry. We optimised the workflowwith particular reference to cell fixation which is crucial for successful laser-based microdissection and also downstream molecular studies. In addition, we defined the minimum number of cells to be isolated and analysed for satisfactory proteome coverage. To set up this workflow, we choose human monocyte-derived macrophages spontaneously differentiated in vitro. These cells, under our culture conditions, show distinct morphotypes, reminiscent of the heterogeneity observed in tissues in various homeostatic and pathological states, e.g. atherosclerosis. This optimised workflow may provide new insights into biology and pathology of heterogeneous cell in culture, particularly when other cell selection approaches are not suitable.

Original languageEnglish
Pages (from-to)2817-2825
Number of pages9
JournalAnalytical and Bioanalytical Chemistry
Volume406
Issue number12
DOIs
Publication statusPublished - 2014

Keywords

  • Heterogeneous cellmorphology
  • Label-freemass spectrometry
  • Laser capture microdissection
  • Proteomics

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Medicine(all)

Fingerprint Dive into the research topics of 'A mass spectrometry-based workflow for the proteomic analysis of in vitro cultured cell subsets isolated by means of laser capture microdissection'. Together they form a unique fingerprint.

  • Cite this