A branched DNA signal amplification assay to quantitate messenger RNA of human uncoupling proteins 1, 2, and 3

Uncoupling proteins (UCP) are inner mitochondrial membrane transporters whi ch dissipate the proton gradient, releasing stored energy as heat. Three su btypes of UCP have been identified so far, The regulation of UCP expression is mainly controlled at the transcriptional level, thus making the measure ment of UCP mRNA beneficial for both diagnosis and research of weight disor ders and diabetes. We have developed an assay using the branched DNA signal amplification assay (bDNA assay) to quantitatively measure the mRNA levels for human UCP1, 2, and 3, UCP-subtype-specific primers were designed for t he assay. RNA transcripts of each UCP generated by in vitro transcription w ere used to validate the specificity and sensitivity of the assay, The quan titative measurement of UCP mRNA was further demonstrated with cultured cel ls and human tissue. A comprehensive survey of UCP expression from 17 human tissues measured by the newly developed assay is provided. The method desc ribed here offers a rapid, sensitive, specific, and quantitative assay for measurement of human UCP mRNA (C) 2000 Academic Press.