Measurement of DNA interstrand crosslinking in individual cells using the Single Cell Gel Electrophoresis (Comet) assay

Methods Mol Biol. 2010;613:267-82. doi: 10.1007/978-1-60327-418-0_17.

Abstract

The Single Cell Gel Electrophoresis (Comet) assay, originally developed to allow visualisation of DNA strand break damage in individual cells, has been adapted to measure DNA interstrand cross-links. DNA interstrand cross-links are formed in cells by a number of commonly used cancer chemotherapy agents and are considered to be the critical lesion formed by such agents. This technique allows the analysis of DNA interstrand cross-link formation and repair at a single cell level, requires few cells, allows the determination of heterogeneity of response within a cell population and is sensitive enough to measure DNA interstrand cross-links at pharmacologically relevant doses. The method can be applied to any in vitro or in vivo application where a single cell suspension can be obtained. The method has also become invaluable in studies using human tissue and can be used as a method for pharmacodynamic analysis in early clinical trials.

MeSH terms

  • Ascites / metabolism
  • Cell Line, Tumor
  • Comet Assay / methods*
  • Cross-Linking Reagents / pharmacology*
  • DNA / chemistry*
  • DNA / genetics
  • DNA / metabolism*
  • DNA Damage
  • Humans
  • Lymphocytes / drug effects
  • Lymphocytes / metabolism
  • Melphalan / pharmacology
  • Staining and Labeling

Substances

  • Cross-Linking Reagents
  • DNA
  • Melphalan