Later ELISA and its various modifications were applied for detection, typing, and strain differentiation of FMDV isolates with better sensitivity than CFT and, the results were comparable to that obtained with MNT [25, 125, [128] [129] [130] [131] [132] [133] [134] . Liquid phase blocking ELISA using bovine convalescent sera for characterization of field isolates was done and result tallied with conventional VNT [135] [136] [137] . ELISA results were much more reproducible than those obtained with VNT and are not influenced by variations in tissue culture susceptibility. At the FAO/WRL for FMD, the preferred procedure for the detection of FMDV antigen and identification of viral serotypes is ELISA [26, 138] . The results can be obtained within 3-4 hours after sample is received by the laboratory; a negative sample is confirmed by inoculation of sample into sensitive cultures followed by confirmation of the virus serotype by ELISA. Indirect ELISA was initially used for detection of FMDV antigen in infected cell culture fluid, mice carcass, and cattle tongue as well as antibodies in sera samples [129, 133] . Later a sandwich ELISA was used for subtype analysis of FMDV isolates [134] . Subsequently, a sandwich ELISA was developed for detection and typing of FMDV directly from field materials [139] . Although ELISA is far finer to CFT, a large number of samples failed to give positive results and such negative sample has to be confirmed by inoculation of sample into sensitive cultures [140] followed by confirmation of the virus serotype by ELISA taking 4 more days, a time frame compatible with the need to rapidly detect disease and initiate and appropriate disease control strategy. As a consequence, there is a need of an alternative assay system that allows more rapid confirmation of clinical diagnosis with more sensitivity and this has resulted in development of polymerase chain reaction (PCR) or the more recent real-time PCR.