Quantitation of mRNA for the beta-adrenoceptor gene in human mononuclear leucocytes by in situ hybridization with fluorochrome labeled cloned DNA.
Determination of the transcriptional activity of the beta-adrenoceptor (beta-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 to adhesive glass slides. After incubation with fluorochrome-labeled cloned hamster beta AR-DNA probes the amount of DNA-mRNA hybrids formed in the cytoplasm of individual cells was analyzed by microfluorimetry of specifically bound fluorochrome. In nine of ten normal subjects studied so far a fraction of beta AR-mRNA positive MNL could be detected. The proportion of positive cells as well as their signal intensity revealed a high interindividual variance.
Liebl
B
B
Department of Internal Medicine, University of Munich, FRG.
Pachmann
K
K
Römer
U
U
Haen
E
E
Hallek
M
M
Emmerich
B
B
D000293:Adolescent; D000328:Adult; D003001:Cloning, Molecular; D015342:DNA Probes; D005260:Female; D016650:Fluorescein-5-isothiocyanate; D005452:Fluoresceins; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000737:chemistry*; D008297:Male; D008875:Middle Aged; D009693:Nucleic Acid Hybridization; D012333:RNA, Messenger / Q000032:analysis* / Q000235:genetics; D011943:Receptors, Adrenergic, beta / Q000235:genetics*; D013861:Thiocyanates; D014158:Transcription, Genetic / Q000235:genetics
D015342:DNA Probes; D005452:Fluoresceins; D012333:RNA, Messenger; D011943:Receptors, Adrenergic, beta; D013861:Thiocyanates; D016650:Fluorescein-5-isothiocyanate