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中国精品科技期刊2020
黄登宇, 高文静, 黄种乾, 高丽霞, 冯敏, 杨秀松. 呋喃唑酮代谢物间接竞争化学发光酶免疫法的建立[J]. 食品工业科技, 2017, (09): 294-299. DOI: 10.13386/j.issn1002-0306.2017.09.048
引用本文: 黄登宇, 高文静, 黄种乾, 高丽霞, 冯敏, 杨秀松. 呋喃唑酮代谢物间接竞争化学发光酶免疫法的建立[J]. 食品工业科技, 2017, (09): 294-299. DOI: 10.13386/j.issn1002-0306.2017.09.048
HUANG Deng-yu, GAO Wen-jing, HUANG Zhong-qian, GAO Li-xia, FENG Min, YANG Xiu-song. Synthesis of furazolidone metabolite artificial antigen and development of indirect competitive chemiluminescence enzyme immunoassay[J]. Science and Technology of Food Industry, 2017, (09): 294-299. DOI: 10.13386/j.issn1002-0306.2017.09.048
Citation: HUANG Deng-yu, GAO Wen-jing, HUANG Zhong-qian, GAO Li-xia, FENG Min, YANG Xiu-song. Synthesis of furazolidone metabolite artificial antigen and development of indirect competitive chemiluminescence enzyme immunoassay[J]. Science and Technology of Food Industry, 2017, (09): 294-299. DOI: 10.13386/j.issn1002-0306.2017.09.048

呋喃唑酮代谢物间接竞争化学发光酶免疫法的建立

Synthesis of furazolidone metabolite artificial antigen and development of indirect competitive chemiluminescence enzyme immunoassay

  • 摘要: 建立快速检测动物源性食品中呋喃唑酮代谢物AOZ的间接竞争化学发光酶免疫法。利用对醛基苯甲酸(4-CBA)将AOZ衍生化为CPAOZ,利用活化酯法将CPAOZ与卵清蛋白(OVA)偶联,生成完全抗原CPAOZ-OVA。对包被原与抗体的最佳稀释倍数、封闭液浓度、竞争时间、酶标二抗稀释倍数进行优化,建立标准曲线,同时对方法的特异性进行评价。结果表明:包被抗原最佳稀释倍数为2000倍,抗体的最佳稀释倍数为20000倍、封闭液为2%脱脂乳粉、竞争时间为2 h、二抗最佳稀释倍数为4000倍;线性范围是0.0757.396 ng/m L,半抑制浓度IC50为0.74 ng/m L,对呋喃唑酮原药的交叉反应率为23.4%,与其他结构类似物及衍生化试剂的交叉反应率均小于0.1%。该方法灵敏度高、特异性好,可为监管部门对于快速检测动物源性食品中AOZ提供依据。 

     

    Abstract: An indirect competitive chemiluminescence enzyme immunoassay ( ic-CLEIA) was used for determination and identification of the furazolidone metabolite in animal-derived food. AOZ can be translated into hapten CPAOZ by derivative reaction.CPAOZ-OVA, as the coating antigen, was prepared by activated ester method.After optimizing the dilution multiple of coating antigen and anti-body, blocking buffer, competitive reaction time and the dilution multiple of second antibody, a standard curve was established, and finally the specificity of the method was evaluated.The results indicated that, the optimum dilution multiple of coating antigen was 2000-fold, the anti-body was 20000-fold.Blocking buffer was dried skim milk at the concentration of 2%.The competitive reaction time was 2 h, and the ideal dilution multiple of secondary antibody was 4000-fold.The linearity range was 0.0757.396 ng/m L, and half-inhibitory concentration ( IC50) value was 0.74 ng/m L.The crossreactivity value between AOZ and furazolidone original drug was 23.4%, and cross reactivity value with other analogues and derivatives was < 0.1%. The method with demonstrated excellent sensitivity and specificity can provide the basis for rapid detection of AOZ content in animal-derived food for supervision department.

     

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