半乳糖-1-磷酸尿苷酰转移酶荧光定量测定及保存条件对酶活性的影响  

Quantitive fluorometric assay for GALT and the influence of different storage condition on activity

作　　者：常会波[1] 王立文[1] 朱彦丽[1] 李媛媛[1] 刘哲伟[1] 张霆[1] 吴建新[1] 

机构地区：[1]首都儿科研究所,北京100020

出　　处：《中国优生与遗传杂志》2010年第6期18-19,26,共3页Chinese Journal of Birth Health & Heredity

基　　金：国家自然科学基金项目(30671156);北京市自然科学基金项目(7042017);北京市优秀人才培养资助项目(20081D0303200106)

摘　　要：目的探讨荧光定量测定导致经典型半乳糖血症的半乳糖-1-磷酸尿苷酰转移酶(GALT)的方法,以及不同保存条件对酶活性的影响。方法选取来本院就诊的半乳糖血症患儿及其父母为研究对象,随机选取健康体检儿童作为对照,采集肝素抗凝血,用定量荧光法测定GALT活性,并观察不同保存温度和不同时间的GALT活性的变化。结果本方法表明酶反应1hr时可以分辨出完全失活型和杂合子,诊断出75%的半乳糖血症患儿,并检测出4例杂合子。同时发现随着时间的推移GALT活性在下降,室温影响最大。结论本方法简单,准确,快捷,费用低廉适合于半乳糖血症的诊断和杂合子的检出,但要注意高温和保存时间对样品的影响。本法也适合于新生儿大规模筛查和孕妇杂合子的检出。Objective ： To develop the quantitive fluorometric assay for galactose - 1 - phosphate uridyltransferase （GALT） activity in blood and dried blood spot specimens on filter paper, and investigate the effect of GALT activity in different storage conditions. Methods ： The fluorometric assay was used to detect GALT activity in blood red cell of galactsim patients and their parents, and normal children. Also samples were stored as whole blood and dried blood spot at room temperature. 4℃, -20℃, and subsequent assays were performed on days 3, 7 and 14 respectively. Results： Patients with GALT deficiency were distinguished clearly from healthy subjects and heterozygous carriers by quantitive fluorometric assay in one hour. 75% galactosemia patients and 4 heterozygous carriers were detected using this method. Also, increased temperature decreased the stability of GALT activity in both liquid blood and dried blood spots. Conclusions ： The quantitative test can provide precise GALT activity and should take into consideration of the influence of high temperature, duration between sampling and testing. This rapid, simple and reliable assay system should be ＇useful for the large - scale screening of newborns for galactosemia and detect heterozygotes of pregnant women.

关 键 词：定量荧光法 半乳糖-1-磷酸尿苷酰转移酶 存储 

分 类 号：R725.8[医药卫生—儿科]