Quantitation of mRNA for the β-Adrenoceptor Gene in Human Mononuclear Leucoocytes by in situ Hybridization with Fluorochrome Labeled Cloned DNA

Abstract

Determination of the transcriptional activity of the β-adrenoceptor (β-AR) gene in human mononuclear leucocytes (MNL) may provide important information concerning variations in receptor expression in clinical studies. We used a new method with highly fluorochrome-labeled gene. probes for in situ hybridization which allows a rapid detection of mRNA at the cellular level. MNL from venous blood of healthy volunteers were permeabilized, fixed and attached t o adhesive glass slides. After incubation with fluorochrome-labeled cloned hamster OAR-DNA probes the amount of DNA-mRNA hybrids formed in the cytoplasm of individual cells was analyzed by rnicrofluorimetry of specifically bound fluorochrorne. In nine of ten normal subjects studied so far a fraction of OAR-mRNA positive MNL could be detected. The proportion of positive cells as well as their signal intensity revealed a high interindividual variance.