血清glypican 3 ELISA检测法的建立及初步应用

Establishment of antibody-sandwich ELISA for detection of serum glypican 3 and its clinical application

目的建立血清中磷酯酰肌醇蛋白聚糖3(glypican 3,GPC3)的双抗体夹心ELISA检测法,并探讨其应用价值。方法以本室自制抗GPC3单克隆抗体为包被抗体,以兔抗人GPC3抗体为检测抗体,建立检测GPC3的双抗体夹心ELISA法。对建立的方法进行线性范围、灵敏度、精密度、回收率等方法学评价,并用于测定原发性肝癌、肝炎后肝硬化、慢性肝炎患者及体检健康者血清标本。结果该法线性范围为0.2～12.5μg/L,灵敏度为0.101μg/L,批内CV≤5.99%,批间CV≤6.96%,平均回收率为99.9%。原发性肝癌患者血清GPC3浓度5.43(2.28～8.62)μg/L明显高于肝炎后肝硬化、慢性肝炎患者及健康对照组(P均<0.05)。以3.19μg/L为临界值时,诊断肝癌的敏感性和特异性分别为85.4%及98.9%。结论该法灵敏、特异和准确,可用于原发性肝癌的诊断。

Objective To establish an antibody-sandwich ELISA method for detection serum glypican 3（GPC3）.Methods The monoclonal and polyclonal antibody to GPC3 were prepared by ourselves,and a double-antibody-sandwich ELISA method for the detection of serum GPC3 was established.Results The linearity was from 0.2 to 12.5 μg/L.The lowest limit was 0.101 μg/L.The coefficients of variation（CV） of within-run and between-run were less than 5.99% and 6.96% respectively,and the average recovery was 99.9%.Clinical studies showed that the concentration of GPC3 in the sera of patients with primary hepatocarcinoma was 5.43（2.28-8.62） μg/L,which was significantly higher than that in benign liver diseases（P0.05）.When cut-off value was 3.19 μg/L,the sensitivity and specificity of primary hepatocarcinoma diagnosis was 85.4% and 98.9% respectively.Conclusion The established ELISA is sensitive,specific and accurate in this study.The detection of serum GPC3 should be helpful for the diagnosis of primary hepatocarcinoma.