Detection of caprine arthritis--encephalitis virus RNA in macrophages by in situ hybridization using fluorescein-labelled single-stranded RNA probes.

The use of in situ hybridization (ISH) for the detection of caprine arthritis-encephalitis virus (CAEV) RNA with fluorescein-11-UTP-labelled single-stranded RNA probes is described. Three different probes were made by PCR amplification of proviral CAEV DNA (strain 75-G63). The PCR products were cloned into the plasmid pAM-18, and labelled single-stranded RNA probes were synthesized by the use of RNA polymerase. The LTR probe was able to detect viral RNA in CAEV-infected, cultured caprine macrophages, while probes based on the genes for the matrix and transmembrane proteins failed to do so. A few macrophages were positive for CAEV RNA 24 h post infection (p.i.) while most cells were positive 96 h p.i. The use of fluorescein-labelled RNA probes made this method feasible for kinetic in vitro studies of CAEV.