血清酸性磷酸酶测定法的实验研究

目的 建立以 2 ,6 -二氯 - 4-乙酰基磷酸盐 (DCAP -P)为底物的一种新的酸性磷酸酶 (ACP)的测定方法。方法 在ISP半自动生化分析仪上研究测定法的试剂配方与实验参数。结果 本法测定DCAP -P吸收峰为 :32 6nm ;Km =0 .139mmol/L ;340nm时DCAP -P的摩尔消光系数为 14 4 4 0L/(mol .cm) ;缓冲液浓度 0 .1mol/L ,最适条件 :pH 5 .4 ,底物浓度 2 .0mmol/L ,延滞期 30s ,线性反应期 15min ;线性范围为 0～ 1188u/L ;高、低值回收率均在 95 %～ 10 5 % ;高、中、低值批内CV分别为 :2 .6 %、1.9%、1.2 % ,批间CV值分别为 :2 .8%、2 .1%、1.2 % ;血红蛋白 (Hb) <115 .6 2mg/L对测定结果无影响 ;黄疸对测定结果无影响 ;与长征试剂盒相比r =0 .978;试剂在 - 2 0℃可稳定一个月 ;本方法的参考值范围为 9u/L～ 2 6u/L。结论 此方法重复性好 ,线性反应期长 ,线性范围广 。

Objective To establish a new determination method by which to measure acid phosphatase activity (ACP) with 2,6-dichloro-4 acetylphenylphosohoric acid as substrate.Methods With the helped of semiautomatic analyzer, made a research on choposition of reagent of continuous-monitoring procedure and experimental parameters.Results When used this assay, the wave-length of DCAP's absorption at 326nm, and the Km of ACP was 0.139?mmol/L. the Molecular extinction coefficient of DCAP of 340?nm was 14?440?L/(mol.cm) and the concentration of citrate's buffer was 0.1?mol/L. During the analytic process, we found that the optimum pH of enzyme was 5.4 and linear range was 0～1?188 u/L, high and low rates of recovery were between 95%～105% respectively. CVs were 2.6%, 1.9%, 1.2% for within-run and 2.8%, 2.1%, 1.2% for between-run. The time of linear reaction was 900 second. The correlation coefficient between present reagent and Chang Zheng kit was r=0.978, P>0.05, stationary phase of reagent was 4 weeks. Serum ACP was 9?u/L～26?u/L respectively which based on the experiment of 30 health adults.Conclusion The method was practical in its repetition and convenience, save time and wasn't liable to be affected by bilirubin in serum, especially suitable for the used of automatic analyzer, and was also useful to the diagnosis of prostatic disease and in judge the therapeutic effect and prognosis of prostatic cancer.