本研究目的在利用牛血清白蛋白與黃麴毒素B1 之共軛物(bovine serum albumin (BSA) conjugate with aflatoxin B1 (AFB1), BSA-AFB1)，以腹腔免疫ICR 小鼠，來生產 黃麴毒素B1 多株抗體。將高度免疫(hyperimmune)之ICR 小鼠，於兩週後注射0.5mL pristane，再間隔兩週注射106 個NS-1 骨髓癌細胞，待腹水產生收集所產生之腹水及血清，利用Hitrap rProtein A column 來純化其多株抗體，並進一步分析已純化anti-AFB1 之抗體效價 (titer) 及其專一性。以間接型enzyme-linked immunosorbent assay(indirect ELISA)進行anti-aflatoxin B1(anti-AFB1)之效價測定。實驗結果顯示，將純化後抗體稀釋10,000 倍情況下，以BSA 濃度100 µg/mL 可中和抗體中的anti-BSA，而得純anti-AFB1 溶液，測其效價為1:10,000。稀釋10,000 倍抗體，建立三明治型ELISA 之AFB1 檢測系統，可測得AFB1 最低濃度在0.4 ng/mL（AFB1 濃度在0.4~10ng/mL 與其A405 呈線性關係）。最後進行AFB1 之交叉反應，結果顯示與AFB2、AFG1 和AFG2 均無交叉反應。以此系統檢測市售玉米罐頭和花生酥中AFB1 之含量，僅玉米罐頭粒狀部分AFB1 含量約40 ng/g (40 ppb)，超過國家規定之15 ppb，其餘玉米罐頭汁液部分和四種花生酥樣本均符合規定。 The aim of this program was to produce polyclonal antibodies against aflatoxin B1 (AFB1), a kind of mycotoxins. By using BSA-AFB1 (bovine serum albumin conjugate with aflatoxin B1 from Aspergillus flavus) to immune intraperitoneally ICR mouse four to five times every four weeks. Hyperimmune ICR mouse produced polyclonal antibodies after injecting with 0.5 mL pristane and NS-1 myeloma cells two weeks later. The polyclonal antibodies were purified by using Hitrap rProtein A column, then the sensitivity and specificity of polyconal antibodies were determined. The anti-BSA of polyconal antibodies were neutralized by 100 �g/mL BSA when the antibodies mixture were diluted 10,000-fold. The titer of anti-AFB1 was 1:10,000 by indirect enzyme-linked immunosorbent assay (ELISA). By diluting 10,000-fold of anti-AFB1, the sandwish ELISA system was established to measure AFB1 concentration. This system could measure the lowest of AFB1 was 0.4 ng/mL (the linear relationship of AFB1 concentration was from 0.4 to 10 ng/mL.), and had low cross reaction with AFB2, AFG1, and AFG2. The AFB1 concentrations in corn cans and peanut butters obtained from supermarkets were determined.
