双标记前列腺特异性抗原时间分辨荧光免疫分析试剂盒研究

The study on the time resolved fluoroimmunoassay of double labeled prostate specific antigen

目的研制钐（Sm）标记检测总前列腺特异性抗原（tPSA）及铕（Eu）标记检测游离前列腺特异性抗原（fPSA）的双标记时间分辨荧光免疫分析法（TRFIA）试剂盒。方法以抗PSA单克隆抗体101^#包被96孔微孔板，用Sm^3＋标记抗tPSA单克隆抗体102^#、Eu^3＋标记抗fPSA单克隆抗体201^#，采用双抗体夹心法建立t／fPSA双标记TRFIA。标准曲线由TRFIA检测仪自带的Log-Log_B函数处理。结果tPSA批内和批间变异系数（CV）分别为2．38％和3．91％，最低检测下限0．02μg／L，可测范围为0．02～250μg／L，回收率为101．3％；fPSA批内和批间CV分别为3．16％和3．34％，最低检测下限0．05μg／L，可测范围为0．05～250μg／L，回收率为103．2％。t／fPSA TRFIA试剂盒测定健康组、前列腺良性疾病组和前列腺癌组（Pca）的tPSA平均值为1．4μg/L、4．3μg/L、14．2μg/L，fPSA平均值为0．3μg/L、1．5μg/L、6．2μg/L。tPSA3．6μg/L、f／tPSA〈0．21为Pca与良性疾病的鉴别阈值。自制试剂盒与PE公司进口试剂盒同时测定血清样本，二者tPSA与fPSA的相关系数分别为0．99和Q97。结论自制t／fPSA TRFIA试剂盒的各项指标均达到临床检测要求，可替代国外同类产品。

Objective To establish the time-resolved fiuoroimmunoassay （TRFIA） of total prostate specific antigen （tPSA） labeled with Sm^3＋ and free prostate specific antigen （fPSA） labeled with Eu^3＋. Methods The monoclonal antibody （McAb） 101^# was coated on the 96 well plates, anti-tPSA McAb 102^# was labeled with Sm^3＋ and anti-fleA McAb 201^# was labeled with Eu^3＋. The standard curves were given out by Log-Log_B function of the DEFIA instrument. The levels of PSA in sera from patients or healthy volunteers were determined by t/fPSA TRFIA using the autoDELFIA1235 system. Results The measurement ranges of tPSA -TRFIA were 0. 02-250 μg/L and those of fPSA-TRFIA were 0.05-250 μg/L. The within-run and between-run CVs of the tPSA-TRFIA were 2. 38% and 3.91% , respectively, and those of fPSA-TRFIA were 3.16% and 3.34% , respectively. The recovery rates of tPSA-TRFIA and fPSA-TRFIA were 101.3% and 103.2%, respectively. The detection limitations of tPSA and fPSA were 0. 05 μg/L and 0. 02 μg/L, respectively. The average results of health group benign prostate disease patients, prostate cancer patients for tPSA were 1.4 μg/L,4. 3 μg/L, 14.2 μg/L, and those of fPSA were 0. 3 μg/L, 1.5 μg/L ,6. 2 μg/L. The cut-off point condition for benign prostate disease and malignant prostate cancer was tPSA 〈 3.6 μg/L, fleA/ tPSA 〈0. 21. The linear equation of tPSA and fPSA tested with PE and our kits were Y =0. 8641X -0. 0779, Y = 0. 9116X - 0. 0123, whose correlation ratio were 0.99 of tPSA and 0.97 of fPSA. Conclusions The availability of a highly sensitive, reliable, and convenient t/fPSA TRFIA method for quantifying PSA will allow investigations into the possible diagnostic value of analysis in various clinical condition and could be the substitute of import produts.