In situ ligation simplified: using PCR fragments for detection of double-strand DNA breaks in tissue sections

Methods Mol Biol. 2011;682:65-75. doi: 10.1007/978-1-60327-409-8_6.


The simplified in situ ligation procedure is described. All reagents for the assay can be easily obtained in any molecular or cell biology laboratory. The technique uses ligation of double-stranded, PCR-derived DNA fragments labeled with digoxigenin or fluorophores for highly selective detection of apoptotic cells in paraffin-embedded tissue sections. Two types of DNA fragments prepared by PCR are employed. The fragment synthesized by Taq polymerase contains single-base 3' overhangs, whereas the Pfu polymerase-made fragment is blunt ended. Both fragments can be used as specific, sensitive and cost-effective DNA damage probes. After ligation to apoptotic nuclei in tissue sections, they indicate the presence of double-strand DNA breaks with single-base 3' overhangs as well as blunt ends.

MeSH terms

  • Animals
  • DNA Breaks, Double-Stranded*
  • DNA Probes
  • Polymerase Chain Reaction / methods*
  • Tissue Fixation / methods*


  • DNA Probes