In situ hybridization for the detection of chicken anemia virus in formalin-fixed, paraffin-embedded sections

The development of a biotinylated in situ hybridization procedure for the detection of chicken anemia virus (CAV) is described. A double-stranded DNA probe was prepared using polymerase chain reaction and was biotinylated by nick translation. Hybridization conditions included the use of a microwave oven to denature target and probe nucleic acids and a five-step protocol for detection of biotinylated hybrids. In situ hybridization detected CAV nucleic acid in thymus tissue from experimentally infected birds after 6 hours, 3 days, and 7 days fixation time in formalin. However, immunocytochemical detection of CAV antigen was severely impaired if tissue was fixed in formalin for more than 6 hours.