Determination of the protooncogene ets-2 gene transcript in human brain at the atto-gram-level by the use of competitive RT/PCR.

Protooncogenes (PO) play a crucial role for brain biology and pathology. Only the concerted action of protooncogenes enables normal brain development. The reliable and sensitive quantification of brain PO is still holding centre stage in neurobiological research. The aim of our study was therefore the determination of PO in minute amounts of brain areas. For this purpose we decided to apply the most sensitive detection principle of competitive reverse transcriptase polymerase chain reaction using capillary electrophoresis and laser-induced fluorescence detection. We selected the PO ets-2 for our studies as this transcription factor was shown to be involved in neurodegenerative disease. As little as 10 ng of total RNA each were extracted from 5 different regions of human postmortem brain and used in the assay system. Our results revealed that the ets-2 gene transcript was detectable at the atto-gram level in the brain (54.5 +/- 17.7 ag/ 10 ng RNA in the occipital lobe, 34.2 +/- 7.5 in temporal lobe, 40.2 +/- 15.6 in the frontal lobe, 31.4 +/- 15.7 in the cerebellum, and undetectably low in the parietal lobe). This is the first report at this sensitivity level providing neurobiology with a powerful analytical tool.