[The detection of viral nucleic acid sequences in paraffin-embedded subacute sclerosing panencephalitis (SSPE) and progressive multifocal leukoencephalopathy (PML) brain tissues by in situ hybridization]

Abstract

We have been able to detect the viral nucleic acid sequences in formalin-fixed paraffin embedded tissue sections of the brain from SSPE and PML respectively by in situ hybridization using cloned and radiolabelled complementary DNA (cDNA) measles nucleocapsid (NP) and JC virus probes which are gene specific respectively. We have mainly followed in situ hybridization procedure reported by Haase et al, but partly modified the technique including the extensive wash by buffer solution and the elevated concentration of proteinase K. Firstly, to compare with the number of hybridized cells and grain counts, the in situ hybridization were performed in SV 40 infected cells treated with ethanol-acetic acid for 15 minutes or 10% neutral formalin for at least one week to simulate procedure used by pathologists. Similar findings were obtained in both treatment of 1 microgram/ml concentration of proteinase K in ordinary fixed cells and in 100 micrograms/ml in formalin-fixed cells. In addition we investigated the ability of in situ hybridization technique to detect measles in SSPE and papova virus in PML obtained postmortem that were been formalin-fixed paraffin-embedded routinely. In paraffin-embedded SSPE cerebral cortex positive hybridization were obtained in oligodendroglias and neurons using 125I-labelled measles NP probe in 6 days' autoradiographic exposure (Fig. 2 and 3) in our modified technique. Additionally, in PML tissue JC virus nucleic acid sequences were also detected in many oligodendroglias and swollen neurons using 3H-labelled JC virus probe.(ABSTRACT TRUNCATED AT 250 WORDS)