Increased Sensitivity of ELISA for Potato Viruses S, X, and Y by Polystyrene Pretreatments, Additives, and a Modified Assay Procedure

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Abstract
The virus-specific absorbance (A405) of ELISA for potato viruses S, X and Y (PVS, PVX and PVY) was increased an average of 14.8% by pretreating new polystyrene cuvettes with 5.3 M NaOH:EtOH. Sodium diethyldithiocarbamate (0.01 M) in phosphate-buffered saline plus Tween 20 (PBS-T) increased the PVY-specific absorbance by 37.3% over that using a PBS-T control. Alkaline degradation of PVX by 0.2 M diethanolamine, pH 10.5, and 6-12% sodium sulfite increased the PVX-specific absorbance by 20.6% over that using PBS-T alone. Enhanced sensitivity was further characterized by regression analysis of the relationship between log A405 and log/sap dilution. Dilution end points for virus detection were predicted from regression equations and corresponded to increases in the virus-sepcific absorbance using undiluted infected sap. A modified double-antibody sandwich ELISA increased the dilution end point for PVS detection by 30-32%, which was less than that previously reported with other plant viruses.