Epidermal cell kinetics by combining in situ hybridization and immunohistochemistry

Double labelling can serve as a useful tool for providing information about cell kinetics in normal and hyperproliferative tissues in general, and ski n 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 prob e specific for histone 2b, 3, 4 mRNA sequences. Double labelling was perfor med on normal, tape-shipped normal skin and psoriatic skin. The three proli feration markers were also examined by single labelling. The ratio of cells in the S-phase (N-s) and the growth fraction (N-cy) was determined. In nor mal 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 doubl e-labelling technique cannot be used routinely. Therefore, in order to unde rstand the cell kinetic processes better, experiments are recommended to op timize these methods. From a practical point of view and for reasons of spe cificity and sensitivity, we prefer the Fluorescein-labelled oligonucleotid e probe method. (C) 1998 Chapman & Hall.