Evaluation of the enzyme-amplified ELISA for the detection of potato viruses A,M, S,X, Y,and leafroll

Various parameters,e.g. types of microtiter plate for DAS-ELISA (double antibody sandwich-enzyme-linked immunosorbent assay), use of fresh or frozen amplifier solutions for enzyme-amplified-ELISA, and use of sodium diethyldithiocarbamate (NaDIECA) in sample buffer in cocktail-ELISA were evaluated for the detection of potato viruses A, M, S, X, Y and leafroll from potato foliage. Dynatech Immulon immunoplates provided higher readings for all viruses. Fresh amplifier solution in amplifed-ELISA was superior to frozen solutions. Amplified ELISA gave only marginal improvement in the sensitivity over the standard DAS-ELISA. Addition of NaDIECA in sample buffer did not improve the detection of viruses in DAS-, amplified-, or cocktail-ELISA. Cocktail-ELISA can reduce antigen incubation time to as short as 15 min for PVA, PVM and PVX; 1 hr for PVY and PLRV; and 2–4 hr for PVS using pre-coated plates. Although amplified-ELISA is slightly more sensitive than DAS-ELISA for certain potato viruses, it is not suitable for large-scale indexing of potato viruses in Seed Certification Laboratories, in view of the additional steps needed in carrying out this procedure.