Fluorometric enzyme immunoassay for measurement of infectious feline leukemia virus and its neutralization.
A fluorometric enzyme immunoassay using an alkaline phosphatase-conjugated monoclonal antibody was developed to quantitate feline leukemia virus (FeLV) infection. Monoclonal antibodies, directed against the FeLV structural protein p27, were conjugated with alkaline phosphatase using a modified maleimide method. The enzyme immunoassay requires only 4 days to reproducibly measure FeLV production instead of the 12 days required for the commonly used transformation assay using C81 cells. A linear correlation was found between the virion associated reverse transcriptase activity and the amount of intracellular p27 as determined by the fluorometric enzyme immunoassay. An immunocytochemical assay using the same conjugated monoclonal with a different substrate gave visible plaques in infected cell monolayers and was therefore used to titrate FeLV in plaque-forming units. The results obtained by all the procedures followed single hit kinetics for FeLV infection. The fluorometric enzyme immunoassay was adapted to measure FeLV neutralizing antibodies, allowing a sensitive and accurate determination of neutralizing titers.
Kushner
N N
NN
Cambridge BioScience Corporation, Biotechnology Research Park, Worcester, MA 01605.
Riggin
C H
CH
Annunziato
M E
ME
Marciani
D J
DJ
D000818:Animals; D000911:Antibodies, Monoclonal; D002415:Cats; D002460:Cell Line; D005347:Fibroblasts / Q000032:analysis; D005470:Fluorometry / Q000379:methods; D015683:Gene Products, gag; D007124:Immunoenzyme Techniques; D007150:Immunohistochemistry; D005256:Leukemia Virus, Feline / Q000032:analysis* / Q000276:immunology; D009500:Neutralization Tests / Q000379:methods; D012191:Retroviridae Proteins / Q000032:analysis; D010948:Viral Plaque Assay / Q000379:methods
D000911:Antibodies, Monoclonal; D015683:Gene Products, gag; D012191:Retroviridae Proteins