Transient transfection assay of cell death genes

Methods Enzymol. 2000;322:480-92. doi: 10.1016/s0076-6879(00)22044-2.

Abstract

In conclusion, transient transfection is an efficient and powerful method to determine quickly whether a gene has a detrimental effect on cell survival. We have described a variety of assay systems from which to choose. Each system has its own advantages and disadvantages. It is important to back up any experimental conclusion with more than one type of assay if possible. In addition, one must consider the fact that transient transfection can often achieve artificially high levels of a protein product that may be unrealistic in vivo. High levels of certain proteins may have an adverse effect on cell survival even when they have nothing to do with apoptosis in vivo. Thus, we must emphasize here that a transient transfection assay is just the first test to determine the function of a gene.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Apoptosis / drug effects
  • Apoptosis / genetics*
  • Benzimidazoles
  • Cell Nucleus / ultrastructure
  • Cells, Cultured
  • Flow Cytometry / methods
  • Fluorescent Dyes
  • Genes, Reporter
  • Green Fluorescent Proteins
  • HeLa Cells
  • Humans
  • Immunohistochemistry / methods
  • In Situ Nick-End Labeling / methods
  • Luciferases / analysis
  • Luciferases / genetics
  • Luminescent Proteins / analysis
  • Luminescent Proteins / genetics
  • Microscopy, Fluorescence / methods
  • Necrosis
  • Recombinant Proteins / analysis
  • Recombinant Proteins / biosynthesis
  • Transfection / methods*
  • Tumor Necrosis Factor-alpha / pharmacology
  • beta-Galactosidase / analysis
  • beta-Galactosidase / genetics

Substances

  • Benzimidazoles
  • Fluorescent Dyes
  • Luminescent Proteins
  • Recombinant Proteins
  • Tumor Necrosis Factor-alpha
  • Green Fluorescent Proteins
  • Luciferases
  • beta-Galactosidase
  • bisbenzimide ethoxide trihydrochloride