Application of in situ PCR for the detection of bovine leukaemia virus (BLV) infection in dendritic cell cultures

Abstract

The aim of the study was to develop an in situ PCR (IS-PCR) method for detection of bovine leukemia virus (BLV) in cell cultures. Samples from five BLV positive and five BLV negative cows were collected and dendritic cells (DCs) from blood, bone marrow, spleen, and lymph node were cultured. Cultures prepared from healthy animals were infected with BLV. After two weeks, the cells were tested by nested PCR and IS-PCR for the presence of proviral DNA. As a positive control adherent cell line permanently infected with BLV was used. BLV was successfully detected by IS-PCR in DCs from naturally infected cattle and DCs infected in vitro. In control, non-infected DCs, the results of the reaction were negative. The results of provirus detection by IS-PCR were similar with these performed with nested PCR. Additionally, IS-PCR provides many advantages, like specific localisation of infection and smaller number of cells needed as template for PCR.