基于胆汁泡相糖蛋白特征而研制诊断胆结石ELISA药盒的可行性分析

目的:分析胆汁泡相糖蛋白的特征并评估其ELISA药盒临床应用的可行性。方法:应用凝集素探针介导的点印迹法分析泡蛋白糖链的组成与结构,利用酶解方法测定泡蛋白各组分的构成,并分析其氨基酸组成。利用Westernblot法分析胆汁与血清泡蛋白的同源性,根据ELISAkit测定泡蛋白的含量分布,并以ROC曲线判别ELISAkit的临床应用价值。结果:化学分析泡蛋白由多肽和寡糖组成,其中多肽分子量约为21u,糖链约占总蛋白含量的37.3%,为核心岩藻糖丰富的多天线复杂型聚糖。泡蛋白具胆汁与血清分布两相性,胆固醇性结石组胆囊胆汁和血清33.5u泡蛋白含量分别为(213.4±70.1)μg/ml、(179.8±97.9)μg/ml,明显高于胆色素性结石症组和正常对照组(P<0.05),而后两者间差异无显著性(P>0.05)。ELISAkit判断胆固醇性结石的胆汁相敏感度和特异度分别为85%和90%,而血清相为61.7%和93.3%。结论:胆汁泡蛋白是一种新认识的胆结石促成核因子,其在不同人群中具有表达谱的异质性,ELISAkit辅助诊断胆石症具有一定的临床应用价值,但尚需更多病例的积累和验证。

The feasibility to develop an ELISA kit based on qualitative and quantitative analyses of vesicular protein in human bile

Objective To analyze the characteristics of "vesicular"protein and to assess the feasibility of developing an ELISA kit for the diagnosis of cholesterol gallstones. Methods The purified 33.5 u "vesicular" protein was detected by lectin,labeled probe and it was deglycosylated by enzymatic approaches to determine the molecular weight of the protein component alone; the type of linkage between the constituent polypeptides and the carbohydrate components was studied also. Furthermore, the distribution of the protein in gallbladder bile and in the serum of patients with cholesterol stone, pigment stone and controls were examined and then analyzed by receiver operating characteristic curve(ROC). Results The 33.5 u "vesicular" protein was composed of polypeptides and an extensively glycosylated (37.3%) monomer. The mean concentration of the 33.5 u "vesicular" protein was significantly greater in cholesterol stone patients than in pigment stone patients and in the controls, both in bile and serum samples. The diagnosis sensitivity and specificity of the ELISA kit for cholesterol stone patients were 85% and 90% respectively in the bile phase, and 61.7% and 93.3% respectively in the serum samples. Conclusions The "vesicular" protein is a novel pro,mucleation factor of the human bile, and distributes unevenly in different patient groups. The ELISA kit recommended in this paper could be a useful mains for the diagnosis of cholesterol gallstone(s).