Screening of zearalenone-producing strains by ELISA method

ELISA법에 의한 zearalenone 생성균주의 검색

ELISA method was applied for the screening of zearalenone producing strains. The developed ELISA was as follow: $125\;{\mu}l$ of diluted solution (1 : 500) of antibody was added to each microtiter well and incubated overnight at $40^{\circ}C$. For direct competitive ELISA, samples and zearalenone-peroxidase conjugate were mixed in a 1 : 1 ratio, and a $100\;{\mu}l$ of aliquot was then added to antisera-coated wells. Plates were incubated for 30 minutes at $37^{\circ}C$, and wells washed 6 times, and $100\;{\mu}l$ of ABTS substrates was added. Plates were incubated for antother 15 minutes at $37^{\circ}C$, and $100\;{\mu}l$ of stopping reagent was added to the wells and absorbance was recorded at 410nm on ELISA Reader. Among 19 strains showed zearalenone-producing ability by ELISA, 3 strains (R-5, C-46, S-134) produced more than 50 ng/ml of zearalenone.

ELISA법을 zearalenone 생성균주 검색에 응용하였다. 먼저 zearalenone에 대한 항체를 500배 희석한 후 microtiter well에 $125\;{\mu}l$씩 주입하여 $40^{\circ}C$에서 overnight시켜 coating하고, 시료용액과 enzyme을 $37^{\circ}C$에서 30분간 반응시켰다. 배양후 washing buffer로 6회 세척한 다음 plate에 2,2'-azino-di-3-ethyl-benzthiazoline sulfonic acid(ABTS) 용액을 $100\;{\mu}l$씩 첨가하여 15분간 발색시킨 다음 반응 정지액 $100\;{\mu}l$씩을 가해 반응을 정지시키고 ELISA Reader로서 흡광도(410 nm)를 측정하였다. 그 결과 zearalenone 생성이 확인된 19 균주 중 분리균 R-5, C-46 및 S-134가 50 ng/ml 이상의 zearalenone을 생성하였다.