尿液总睾酮测定的新方法

A New Method of Total Urinary Testosterone Determination with a Highly Sensitive Time-resolved Fluorescence Immunoassay

目的:用高灵敏时间分辨荧光免疫分析(TrFIA)技术建立尿总睾酮快速准确检测方法。方法:将人睾酮包被微孔板,加入高亲和力的羊抗人睾酮抗体,然后用Eu3+-DTTA标记的猴抗羊抗体作检测抗体,建立基于竞争法反应原理的TrFIA。按常规方法对该法的特异性、灵敏度、准确度、精密度及线性等指标进行考核。用该法检测260例(男137,女123)正常成人尿液总睾酮含量,并与放射免疫分析(Radioimmunoassay,RIA)方法进行比较。结果:TrFIA检测尿总睾酮的灵敏度为5.1pmol/L,特异性、精密度、准确度、线性等指标都达到临床检测试剂盒的要求,且在4℃可稳定保存半年以上。用该法测定正常成人尿液中总睾酮浓度,男性为(40～233.82)nmol/L(5%～95%区间),平均值为(134.26±60.39)nmol/L;女性为(9.44～67.1)nmol/L(5%～95%区间),平均值为(32.18±18.00)nmol/L。TrFIA法检测结果与商用RIAkit检测结果相关系数为0.992,回归方程为:RIA=0.075+0.971TrFIA。结论:TrFIA测定尿总睾酮快速且准确,可用于临床尿总睾酮常规检测。该法的最大优点是样本无需作前期处理,便可直接测定。

Objective To develop a method of highly sensitive and accurate time - resolved fluorescence immtmoassay （TrFIA） for total urinary testosterone determination in our laboratory. Methods High affinity anti - testosterone antibody and Eu^3＋ labeled monkey anti - sheep IgG as tracer were used to develop the TrFIA method. The assay was evaluated for specificity, sensitivity, recovery rate, precision and dilution linearity on 260 urine specimens and the results compared with these from conventional RIA. Results A satisfactory standard curve for testosterone TrFIA was developed with good sensitivity （5.1 pmol/L）. Specificity, recovery rate, precision and dilution linearity studies were determined and all found to be satisfactory. Male urinary total testosterone excretion ranged from 40 to 233.82 nmol/L, which was about 4 times of the value for women （9.44 - 67.1 nmol/L）. The validity of urinary testosterone assay was confirmed by the good correlation between the results obtained by TrFIA （X） and conventional RIA （Y） （ Y = 0. 075 ＋ 0. 971X, r = 0.992）. Conclusion A rapid, reliable, easy to automate, highly sensitive and specific TrFIA type assay for the measurement of total testosterone in urine specimens was developed originally in our laboratory. The advantage of this novel assay is that it does not require an initial extraction step nor involve time consuming procedures such as chromatography.