ABSTRACT
A polyclonal antibody-based sandwich ELISA (s-ELISA) was developed for the detection of bluetongue viruses (BTV) in cell culture lysates and blood samples of sheep infected experimentally. Rabbit antiserum to purified BTV particles and guineapig antiserum to core particles were used as capture antibody and detection antibody respectively. The assay has detected several of the BTV serotypes isolated in India so far. Other common viruses of small ruminants did not cross-react in the assay. The analytical sensitivity of the assay was estimated to be between 10(2.4) and 10(2.6)TCID(50)/ml with different serotypes of BTV. The sensitivity was compared with that of the reverse transcription polymerase chain reaction (RT-PCR) and the latter was found to be at least 100 times more sensitive. In the infected sheep, BTV antigen(s) was detected in blood as early as on 5-day post-infection (dpi) till 35 dpi. The assay may be useful for testing large number of samples in a very short time.
Subject(s)
Antibodies, Viral , Blood/virology , Bluetongue virus/isolation & purification , Bluetongue/diagnosis , Enzyme-Linked Immunosorbent Assay/methods , Animals , Bluetongue virus/immunology , Cell Culture Techniques , Guinea Pigs , India , Rabbits , Sensitivity and Specificity , SheepABSTRACT
A polyclonal antibody-based sandwich enzyme-linked immunosorbent assay (s-ELISA) was developed for the detection of bluetongue virus (BTV). The test used antiserum against BTV and antiserum against the bluetongue (BT) core protein. The antiserum against the virus was used as a capture antibody and the antiserum against the protein was used for detection. In this study, antiserum to recombinant viral protein 7 (rVP7) was used as a detection antibody in place of anti-core antiserum. The assay was used to detect the BT serotypes found in India, namely: 1, 2, 9, 15, 18 and 23. The modified sandwich assay was able to detect BTV serotypes in cell culture supernatants. The use of anti-rVP7 antiserum as the detection antibody avoids the tedious and expensive purification of BTV core particles.
