| Developing Polyclonal Antibody-based Indirect-ELISA to Detect Anthracnose Inocula Prior to Soybean Sprout Rot |
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Soo Bong Park, Young Ji Lim, Jung Han Lee, Ki Soo Han, Sun Cheol Lee, Chang Ki Shim, Jin Ho Kang, Dong Won Bae, Dong Kil Kim, Hee Kyu Kim |
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| Abstract |
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We developed a polyclonal antibody based-ELISA system to monitor inocula accurately and rapidly before onset of anthracnose on soybean sprouts. Titer of mouse antisera against conidia of Colletotrichum gloeosporioides, determined by indirect ELISA, was high enough to be detectable up to x25,600 dilutions. Both PAb1 and PAb2 had the highest level of reactivity to Colletotrichum gloeosporioides. Absorbance readings exceeded 0.15. Sensitivity of PAb to C. gloeosporioides was precise enough to detect spore concentration as low as 500 conidia/well by indirect ELISA. Both antibodies are very sensitive and highly specific to the target pathogen Colletotrichum gloeosporioides, apparently discriminating other unrelated pathogen, or epiphytes. This kit fulfills the requirements for detecting inocula before infection and onset of anthracnose. Our ELISA system should also be feasible to detect C. acutatum (Mungbean sprouts rot) and G. cingulata (C. gleosporioides), (apple, pepper). It was remarkable that absorbance value was not reduced even after 4 consecutive washings (Fig. 4), suggesting that antigenic determinants are on the surface of conidia. Antigenic determinant was characterized by heating and enzyme treatment: Both PAb1 and PAb2 bind to protein epitope that does not contain residue of amino acid, arginine, and lysine, even though more work needs to be done. |
| Key Words:
anthracnose, indirect ELISA, soybean sprout |
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