Epidermal cell kinetics by combining in situ hybridization and immunohistochemistry

Histochem J. 1998 Dec;30(12):869-77. doi: 10.1023/a:1003457709690.

Abstract

Double labelling can serve as a useful tool for providing information about cell kinetics in normal and hyperproliferative tissues in general, and skin in particular. We have developed a double-labelling method that combines immunohistochemistry using the monoclonal antibody MIB1 and non-isotopic in situ hybridization using either a digoxigenin-labelled RNA probe specific for histone 3 mRNA sequences or a Fluorescein-labelled oligonucleotide probe specific for histone 2b, 3, 4 mRNA sequences. Double labelling was performed on normal, tape-stripped normal skin and psoriatic skin. The three proliferation markers were also examined by single labelling. The ratio of cells in the S-phase (Ns) and the growth fraction (Ncy) was determined. In normal skin, psoriatic skin and tape-stripped normal skin after 24 h and after 48 h, we calculated that 15%, 16%, 3% and 12% of growth fraction consisted of cells in the S-phase respectively. The S-phase lasts approximately 10 h, so the cell cycle time in normal and psoriatic skin is approximately 62.5 h. At present, the MIB1/H3 digoxigenin or MIB1/H2b-H3-H4 Fluorescein double-labelling technique cannot be used routinely. Therefore, in order to understand the cell kinetic processes better, experiments are recommended to optimize these methods. From a practical point of view and for reasons of specificity and sensitivity, we prefer the Fluorescein-labelled oligonucleotide probe method.

MeSH terms

  • Cell Division
  • DNA, Complementary
  • Digoxigenin
  • Epidermal Cells*
  • Epidermis / chemistry*
  • Fluorescein-5-isothiocyanate
  • Histones / genetics
  • Humans
  • Immunohistochemistry
  • In Situ Hybridization
  • Ki-67 Antigen / analysis
  • Kinetics
  • Oligonucleotide Probes
  • Skin / chemistry
  • Skin / cytology
  • Skin / pathology
  • Statistics as Topic

Substances

  • DNA, Complementary
  • Histones
  • Ki-67 Antigen
  • Oligonucleotide Probes
  • Fluorescein-5-isothiocyanate
  • Digoxigenin