Determination of teliospore concentration as surface contaminants in Karnal bunt-infected seed samples of wheat using indirect ELISA

The study describes the indirect ELISA method for the determination of teliospore concentration in infected seed lots employing anti-teliospore antiserum generated against intact teliospores of Tilletia indica - a quarantined pathogen. The detection limit is as small as 5-1000 teliospores. The teliospores were extracted by washing seed either one or three times using detergent to ensure complete removal of teliospores present as surface contaminants in wheat samples with varying per cent of Karnal bunt infection. To ensure the concentration of teliospores within the sensitivity range, optimal volume of different extracted samples was determined. The ELISA reactivity was used to extrapolate the number of teliospores present in wheat samples. Forty-nine wheat samples received for certification based on per cent of infection were also subjected to indirect ELISA to determine the sensitivity of the protocol. The samples having 0.05-26% infection were having teliospore concentration in the range of 7×102 to 6.6×104/1000 seeds.