Identification of apoptotic cells by formamide-induced DNA denaturation in condensed chromatin

O. S. Frankfurt, A. Krishan

Research output: Contribution to journalArticle

148 Citations (Scopus)

Abstract

In this article we describe a novel effect of formamide on DNA of apoptotic nuclei and present a method for specific detection of apoptotic cells based on this effect. Our observations show that formamide induces DNA denaturation in apoptotic nuclei but has no such effect on DNA of non-apoptotic cells. Formamide-induced DNA denaturation combined with detection of denatured DNA with a monoclonal antibody (MAb) against single-stranded DNA made it possible to specifically identify the apoptotic cells. This procedure produced intense staining of the condensed chromatin in the apoptotic nuclei. In contrast, necrotic cells from cultures treated with sodium azide, saponin, or hyperthermia did not bind this antibody, demonstrating the specificity of the formamide-MAb assay for the apoptotic cells. However, TUNEL stained 90-100% of necrotic cells in all three models of necrosis. Because the MAb did not stain cells with single- or double-stranded DNA breaks in the absence of apoptosis, we conclude that staining of the apoptotic nuclei is not influenced by DNA breaks and is induced by specific changes in condensed chromatin, such as damage to the DNA-histone interactions. Importantly, the formamide-MAb technique identified apoptotic cells in frozen sections and in histological sections of formalin-fixed, paraffin-embedded tissues.

Original languageEnglish
Pages (from-to)369-378
Number of pages10
JournalJournal of Histochemistry and Cytochemistry
Volume49
Issue number3
StatePublished - Mar 8 2001
Externally publishedYes

Fingerprint

Nucleic Acid Denaturation
Chromatin
Monoclonal Antibodies
DNA
Staining and Labeling
Sodium Azide
DNA Breaks
Antibody Specificity
Double-Stranded DNA Breaks
Single-Stranded DNA
Saponins
In Situ Nick-End Labeling
Frozen Sections
formamide
Paraffin
Histones
Formaldehyde
DNA Damage
Fever
Necrosis

Keywords

  • Apoptosis
  • Chromatin
  • DNA denaturation
  • Formamide
  • Immunohistochemistry
  • Monoclonal antibody
  • Single-stranded DNA
  • TUNEL

ASJC Scopus subject areas

  • Anatomy
  • Cell Biology

Cite this

Identification of apoptotic cells by formamide-induced DNA denaturation in condensed chromatin. / Frankfurt, O. S.; Krishan, A.

In: Journal of Histochemistry and Cytochemistry, Vol. 49, No. 3, 08.03.2001, p. 369-378.

Research output: Contribution to journalArticle

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