AFP时间分辨荧光免疫分析试剂盒的研制及其临床应用

Preparation of a Time-Resolved Fluoroimmunoassay kit for AFP Detection in This Laboratory and Its Clinical Application

目的:研制AFP时间分辨荧光免疫分析(TrFIA)试剂盒。方法:采用双抗体夹心法建立AFP-Tr-FIA试剂盒,对反应条件进行优化,并对试剂盒的各项指标进行评价。结果:抗体包被浓度确定为5μg/m l,铕标抗体最佳稀释比为1 50。试剂盒的线性范围为1ng/m l^1210ng/m l,灵敏度为0.41ng/m l,准确度高,批内和批间变异系数分别5.1%~8.8%,6.7%~11.9%。与CA199、CA125、CEA和白蛋白无交叉反应。破坏试验表明试剂在37℃可稳定7d。收集50例肝癌患者和370例健康人血清,用本试剂盒测得肝癌患者血清AFP浓度(557.3ng/m l±322.1ng/m l)显著高于健康人(6.5ng/m l±5.4ng/m l)(P<0.001)。370例正常人血清标本测试该试剂盒的正常参考范围为(0~12.0)ng/m l。本试剂盒检测结果与商用W allac AFP试剂盒检测结果相关系数为0.9988。结论:试剂盒各项指标达到规定的要求,可用于临床血清AFP检测。

Objective To prepare a time - resolved fluoroimmunoassay （TrFIA） kit for detecting AFP in clinical use. Methods A double antibody sandwich TrFIA kit for AFP detection was developed in our laboratory using monoclonal antibody of anti - AFP. Results This AFP - TrFIA kit was capable of detecting AFP within the range of 1 ～1210ng/ml with the sensitivity of O. 41ng/ml and without cross - reactivity with CA199, CA125, CEA, or albumin. The intra - and inter - assay coefficients of variation were 5.1%～ 8.8% and 6.7 % ～11.9%, respectively. The prepared AFP - TrFIA reagent was stable at 37℃ for 7 days. Serum samples were collected from 50 patients with proven hepatocellular carcinoma and 370 controls. Serum AFP levels were significantly higher in the patients than those in controls （557.3ng/ml± 322. lng/ml vs 6.5ng/ml ±5.4ng/ml, P 〈0. 001 ）. The correlation coefficient of the detection results between this kit and the commercially available Wailac kit for 100 blood samples was 0. 9988. Conclusion The prepared TrFIA kit for AFP detection meets the demand of clinical application with good sensitivity, precision, specificity and accuracy.