Immunolabeling artifacts and the need for live-cell imaging

Ulrike Schnell, Freark Dijk, Klaas A. Sjollema, Ben N. G. Giepmans*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

272 Citations (Scopus)

Abstract

Fluorescent fusion proteins have revolutionized examination of proteins in living cells. Still, studies using these proteins are met with criticism because proteins are modified and ectopically expressed, in contrast to immunofluorescence studies. However, introducing immunoreagents inside cells can cause protein extraction or relocalization, not reflecting the in vivo situation. Here we discuss pitfalls of immunofluorescence labeling that often receive little attention and argue that immunostaining experiments in dead, permeabilized cells should be complemented with live-cell imaging when scrutinizing protein localization.

Original languageEnglish
Pages (from-to)152-158
Number of pages7
JournalNature Methods
Volume9
Issue number2
DOIs
Publication statusPublished - Feb-2012

Keywords

  • ELECTRON-MICROSCOPY
  • MAMMALIAN-CELLS
  • CULTURED-CELLS
  • IMMUNOFLUORESCENCE MICROSCOPY
  • FLUORESCENCE MICROSCOPY
  • DIFFERENTIAL EXTRACTION
  • IMPROVED PRESERVATION
  • MEMBRANE-PROTEINS
  • IN-VIVO
  • FIXATION

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