赭麴毒素A (Ochratoxin A, OTA)為一個廣泛污染飼料和食物的黴菌毒素，它被認為跟巴爾幹半島地區性的腎疾病和上泌尿道的癌症有關。 為了發展更快速且敏感的方法來檢測OTA，本實驗室用免疫OTA-KLH的BALB/c品種老鼠的脾臟細胞與NS-1骨髓瘤細胞融合，篩選得到一個穩定的融合瘤細胞株H9，用來生產抗OTA的單株抗體。 我們用OTA專一性的單株抗體建立了敏感性高的直接競爭型酵素連結免疫吸附分析法和以奈米金粒子為標記物的快速免疫層析試紙分析法。 在cd ELISA中，抑制50%的OTA-HRP與抗體結合所需OTA的濃度(IC50)為0.27 ng/mL。 OTA專一性的快速免疫層析試紙分析法偵測OTA的敏感性可低到5 ng/mL。 由檢測咖啡樣品中OTA含量的結果可看出免疫層析試紙分析法和ELISA的測試結果有很好的一致性，且結果可直接目測並在少於10分鐘內快速完成。 由本研究所建立的以OTA單株抗體為主的cdELISA和免疫層析試紙分析法可用來快速簡便的監控咖啡樣品中的OTA且樣品不需經過純化。多摩酸(Domoic acid, DA)為一種具有神經刺激性的小分子毒素，主要是由矽藻Pseudonitzschia pungens所產生的二級代謝物，人類誤食被DA污染之貝類後，主要會造成短暫性記憶喪失型中毒，一般稱為Amnesic shellfish poisoning (ASP)症狀，高劑量中毒時甚至會導致死亡。 為了發展更快速且敏感的方法來檢測DA，我們也生產了DA的單株抗體9F1F11，是由免疫DA-KLH的BALB/c品種老鼠的脾臟細胞與NS-1骨髓瘤細胞融合，篩選得到穩定的融合瘤細胞株9F1F11所分泌。 我們用DA專一性的單株抗體建立了敏感性高的直接競爭型酵素連結免疫吸附分析法和以奈米金粒子為標記物的快速免疫層析試紙分析法。 在cd ELISA中，抑制50%的DA-HRP與抗體結合所需DA的濃度(IC50)為0.58 ng/mL。 DA專一性的快速免疫層析試紙分析法偵測DA的敏感性可低到5 ng/mL。 由檢測貽貝樣品中DA含量的結果可看出免疫層析試紙分析法和ELISA的測試結果有很好的一致性，且結果可直接目測並在少於10分鐘內快速完成。
Ochratoxin A (OTA) is a widespread mycotoxin contaminating feed and food, and is suspected of being the etiological agent of Balkan endemic nephropathy (BEN) and its associated urinary tract cancers. To develop more rapid and sensitive methods for detection of OTA, monoclonal antibodies (mAb) against OTA were produced from a stable hybridoma cell line, H9, generated by the fusion of P3/NS-1/1-AG4-1 myeloma cells with spleen cells isolated from BALB/c mouse immunized with OTA-keyhole limpet hemocyanin. The OTA-specific mAb H9 was used for developed a sensitive direct competitive enzyme-linked immunosorbent assay (cdELISA) and a rapid mAb based colloidal gold immunostrip assay. The concentration causing 50% inhibition (IC50) of binding of OTA–horseradish peroxidase with the antibodies by OTA in the cdELISA were found to be 0.27 ng/mL. The detection limit of OTA-specific immunostrip was 5 ng/mL for OTA. The results of immunostrip analysis of OTA in coffee samples were in good agreement with those obtained from cdELISA and the result can be observed directly by the nake eyes, and the detection time could be completed within 10 min. The OTA mAb-based cdELISA and immunostrip assay development in this study is suitable for the simple and rapid screening of OTA in coffee samples without sample cleanup. Domoic acid (DA) is a naturally occurring neuroexcitatory toxin produced primarily by the marine diatom pseudonitzschia pungens. Human ingested DA contaminated shellfish leads to amnesic shellfish poisoning (ASP), which even cause death at high dose of DA. To develop more rapid and sensitive methods for detection of DA, monoclonal antibodies (mAb) against DA were also development, which was produced from a stable hybridoma cell line, 9F1F11, generated by the fusion of P3/NS-1/1-AG4-1 myeloma cells with spleen cells isolated from BALB/c mouse immunized with DA-keyhole limpet hemocyanin. The DA-specific mAb 9F1F11 were used for developed a sensitive cdELISA and a rapid mAb based colloidal gold immunostrip assay. The concentration causing 50% inhibition (IC50) of binding of DA–horseradish peroxidase with the antibodies by DA in the cdELISA were found to be 0.58 ng/mL. The detection limit of the DA-specific immunostrip was 5 ng/mL for DA. The results of immunostrip analysis of DA in blue mussel samples were in good agreement with those obtained from cdELISA and the result can be observed directly by the nake eyes, and the detection time could be completed within 10 min.
