USE OF PROTEINASE-K FOR RT-PCR OF CYTOKINE MESSENGER-RNA IN FORMALIN-FIXED TISSUE

Fresh tissue from cases of sudden infant death syndrome is becoming in creasingly scarce and therefore researchers interesting in studying th e aetiology of this syndrome have had to resort to archival tissue, us ually in the form of paraffin wax sections. A simple method for isolat ing mRNA from formalin fixed, paraffin wax embedded material of suffic ient purity for reverse transcription (RT)-PCR is described. Proteinas e K treatment of formalin fixed, wax embedded tissue followed by RNA S TAT-60 extraction was successful in isolating mRNA suitable for RT-PCR . Interleukin (IL)-1 alpha, IL-6 and tumour necrosis factor (TNF) tran scripts were amplified successfully from heart, but not thyroid, kidne y or liver tissue, of a patient who died following rejection of a tran splanted heart, and IL-1 alpha, but not IL-6 or TNF, transcripts from lung tissue of a six month old baby who died of viral pneumonia. Trans cripts of a housekeeping gene were detected in all tissues. This metho d should be useful for examining gene expression in archival material.