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Title: Amplification of specific mRNA from a single human renal glomerulus, with an approach to the separation of epithelial cell mRNA.
Authors: Bicknell, GR
Shaw, JA
Pringle, JH
Furness, PN
First Published: Oct-1996
Citation: J PATHOL, 1996, 180 (2), pp. 188-193
Abstract: A method has been developed by which single human glomeruli may be plucked from fresh renal biopsies under direct vision, followed by separation of mRNA using oligo-dT-linked paramagnetic beads. The mRNA was amplified by reverse transcription and polymerase chain reaction (RT-PCR). Primers for a variety of human and rat proteins have been developed. The quantity of the amplified cDNA was measured by an enzyme-linked immuno-sorbent assay (ELISA), where biotinylated forward strands of DNA were captured, probed with a fluorescein-conjugated DNA oligomer, and then assayed with an enzyme-linked anti-fluorescein antibody. The cDNA-linked beads are reported to be stable and can be reused with different primer sets, thus forming a "bank' of samples from cases with defined glomerular disorders, which can be used to address new questions as they arise. Using rat glomeruli, a method has been devised which permits at least partial separation of epithelial cell mRNA from mesangial and endothelial cell mRNA.
DOI Link: 10.1002/(SICI)1096-9896(199610)180:2<188::AID-PATH639>3.0.CO;2-K
ISSN: 0022-3417
Type: Journal Article
Appears in Collections:Published Articles, Dept. of Cancer Studies and Molecular Medicine

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