Assay of arbovirus neutralizing antibody by micro methods. Trans R Soc Trop Med Hyg 63: 446–455

Extensive serological surveys require simple, reproducible and specific antibody assay systems. The haemagglutination-inhibition (HI) test (CLARKE and CASALS, 1958) has often been used for arbovirus erological surveys, but it may be complicated by nonspecific reactions despite kaolin or acetone extraction of thesera (HOLDEN, MUTH and SHRINER, 1966). Kaolin extraction may remove 40-60 % of the specific immuno-globulins from the sera (MANN et al., 1967). H I antibody is cross reactive within arbovirus groups and interpretation of results in terms of specific viruses is frequently difficult. To confirm the presence of antibody, sera positive by HI test must, therefore, be retested by virus neutralization. Neutralization tests are therefore needed which are as efficient and economic of materials as the HI test, but avoid the disadvantages of the HI test. We describe two neutralization tests developed for antibody assays with arboviruses in Thailand, including Japanese encephalitis (JEV), dengue (DV) types 1-4, chikungunya (CV), Sindbis (SV), Wesselsbron (WV) and Batai (BV) viruses. Materials and methods Cell cultures and med ium * o BHK-21 cells were grown in medium (M) 199 with 5 ~o foetal bovine serum (FBS) inactivated at 56°C. for 1 hour, and 0"07~o NaHCO3, 200 units penicillin and'200 ~g. streptomycin. The propagation of LLC-MK2 cells has been described previously (RusSELL et al., 1967)