新型肝癌标志物Glypican-3时间分辨荧光免疫法的建立

目的建立磷脂酰肌醇蛋白聚糖3（GPC3）TRFIA，探讨血清GPC3检测在肝癌诊断中的价值。方法以抗GPC3单克隆抗体7C8包板，铕（Eu^3＋）标记GP9单克隆抗体进行检测，建立了双抗夹心TRHA，并对41例原发性肝细胞肝癌及44例慢性乙型肝炎患者血清GPC3进行定量分析。采用放射免疫法检测AFP。采用SPSS12．0软件进行非参数秩和检验。结果GPC3双抗夹心TRFIA线性可测范围为3．125～600ug／L，检测灵敏度为2．06ug／L，批内和批间CV分别为12．25％和12．91％。41例肝癌患者血清中GPC3平均质量浓度为（86．68±110．39）ug／L（中位数56．98ug/L）；而44例肝炎患者血清GPC3浓度为（14．77±29．48）ug／L（中位数2．20ug／L），低于肝细胞肝癌组（Wilcoxon W：1335．00，Z=-4．99，P〈0．001）。根据ROC曲线分析结果，以42．94ug／L为诊断界值时，GPC3诊断肝癌的灵敏度和特异性分别为58．5％（24／41）和95．5％（42／44）。联合GPC3检测，能将AFP诊断肝癌的灵敏度从46．3％（19／41）提高至78．0％（32／41）。结论建立了检测GPC3的TRFIA新方法；该法与AFP联合检测，可提高诊断HCC的阳性率。

Time resolved fluoroimmunoassay for Glypican 3 and its preliminary application

Objective To establish a time resolved fluoroimmunoassay (TRFIA) method for detecting Glypican 3 (GPC3) and to explore the diagnostic value of serum GPC3 for hepatic carcinoma (HCC). Methods Microplate coated with anti-GPC3 monoclonal antibody 7C8 and GP9 labeled with Eu3+ were used to establish TRFIA kit. The serum concentrations of GPC3 in 41 HCC patients and 44 chronic hepatitis (CH) patients were quantitatively analyzed. AFP was detected by with lowest limit of 2.06 μg/L. The CV of inter and intra assay were 12.25% and 12.91%, respectively. The average serum concentration of GPC3 in HCC patients was (86.68±110.39) μg/L (median: 56.98 μg/L). But in CH patients it was only (14.77±29.48) μg/L, which was significantly lower than that in HCC (Wilcoxon W=1335.00, Z=-4.99, P<0.001). With diagnostic cut-off value set at 42.94 μg/L, the diagnostic sensitivity and specificity of TRFIA GPC3 for HCC were 58.5% (24/41) and 95.5%(42/44) respectively. The diagnostic sensitivity of AFP was 46.3% (19/41) in 41 HCC patients, and was raised to 78.0% (32/41) when combined with GPC3. Conclusions Serum GPC3 assay by TRFIA is established and it could increase the diagnostic sensitivity for HCC when combined with AFP.