Purification of a new low molecular weight protein and its clinical application

从尿液中分离出一种新的低分子量蛋白(LMWP)(蛋白1)，并对其理化性质进行了鉴定，测定了其部分氨基酸序列，从而制备了抗该蛋白的单抗。在建立了该蛋白的酶联免疫吸附分析方法后，该蛋白在实验室医学上的意义得到了广泛的研究。1)一种新的LMWP(蛋白1)的纯化和性质：通过检测粗提物与各种抗血清(抗体)的反应活性，在SDS-PAGE上检测到一种未知的蛋白为蛋白1。因此，本项目被修改为使用抗体作为探针来分离蛋白1。建立的纯化程序包括硫酸铵沉淀、ConA亲和、凝胶过滤、抗视黄醇结合蛋白抗体偶联亲和和阴离子交换柱。经SDS-PAGE鉴定其纯度为14 kDa，p1为4.7。测定了部分氨基酸序列，证明与人肺Clara细胞10 kDa分泌型蛋白完全一致。2)制备抗人肺Clara细胞10 kDa分泌型蛋白的单抗。3)ELISA方法的建立：以纯化物为标准，初步建立了一种可靠、准确的检测方法。4)临床应用：蛋白1在各种体液中均有分布，尤其是正常成人的血清值在2~20µg/L之间，尿值存在明显的性别差异，男性高于女性。另一项广泛的研究最终发现了另一个合成部位--前列腺癌。对这种蛋白质的进一步研究正在从根本上和临床上进行。

The authors have isolated a new low molecular weight protein (LMWP) (protein 1) from urine, characterized its physicochemical properties, determine it partial amino acid sequences, and thus prepared a monoclonal antibody against this protein. After establishing an enzyme-linked immunosorbent assay (ELISA) for this protein, the significance of this protein in laboratory medicine has been extensively studied.1) Purification and characterization of a new LMWP (protein 1):An unknown protein detected on SDS-PAGE so far proved to be protein 1 after checking the reactivity of rough purified fraction with various antisera (antibody). The present project is thus modified to isolate protein 1 using the antibody as a probe. The established purification procedure comprised of ammonium sulfate precipitation, ConA affinity, gel-filtration, anti-retinol binding protein antibody coupled affinity, and an anion exchange column. Purity was confirmed by SDS-PAGE and its molecular weight was 14kDa and p1 was 4.7. Partial amino acid sequences were determined which proved to be identical with human lung Clara cell 10 kDa secretary protein.2) Preparation of a monoclonal antibody.3) Establishment of an ELISA:A reliable precise assay was initially developed using purified substance as standard and its monoclonal anbitody.4) Clinical application:Protein 1 is distributed in various body fluids, in particular, serum value ranged in normal adults ranged from 2 to 20 mug/l and urine value was varied with marked sex-differences being higher in male than in female. Another extensive studies finally spot another synthetic sites as prostate. Further studies on this protein was both fundamentally and clinically in progress.