一种利用碱性pH条件下免疫反应性增强的血清鸟氨酸氨甲酰基转移酶敏感酶联免疫吸附测定法。

A sensitive ELISA for serum ornithine carbamoyltransferase utilizing the enhancement of immunoreactivity at alkaline pH.

作者信息

Murayama Hiroshi, Igarashi Makoto, Mori Masataka, Fukuda Yoshihiro, Ikemoto Masaki, Nagata Atsuo

机构信息

Immunology Laboratory, Diagnostics Department, YAMASA Corporation, 2-10-1 Araoi-cho, Choshi, Chiba, 288-0056, Japan.

出版信息

Clin Chim Acta. 2006 Jun;368(1-2):125-30. doi: 10.1016/j.cca.2005.12.017. Epub 2006 Jan 30.

DOI:10.1016/j.cca.2005.12.017

PMID:16445902

Abstract

BACKGROUND

We developed a new enzyme-linked immunosorbent assay (ELISA) for ornithine carbamoyltransferase (OCT) and evaluated its usefulness.

METHODS

Recombinant human OCT expressed in E. coli was used as an antigen to obtain the monoclonal antibodies for this assay.

RESULTS

The reactivity of the antibodies to native OCT as well as recombinant OCT was enhanced at alkaline pH (8.5-10), and the assay's sensitivity was markedly improved. The antibodies react identically with native and recombinant OCT at pH 9.4. The dilution test showed a good linearity between dilution ratios and the concentrations. Different concentrations of OCT added were recovered on average at 90.4%. There was a good correlation between OCT protein levels in the ELISA and OCT enzyme activities (r=0.987, p<0.0001). A significant difference in the serum level of OCT was observed between chronic hepatitis patients (110.7+/-80 ng/ml) and healthy subjects (34.4+/-20.7 ng/ml) (p<0.0001). The serum levels of OCT between sexes differed significantly in the healthy subjects (p<0.0001).

CONCLUSIONS

Our newly established ELISA for OCT using monoclonal antibodies is sensitive enough for clinical application.

摘要

背景

我们开发了一种新的鸟氨酸氨基甲酰转移酶（OCT）酶联免疫吸附测定法（ELISA）并评估了其效用。

方法

以在大肠杆菌中表达的重组人OCT作为抗原，获得用于该测定的单克隆抗体。

结果

抗体对天然OCT以及重组OCT的反应性在碱性pH（8.5 - 10）下增强，并且该测定的灵敏度显著提高。抗体在pH 9.4时对天然OCT和重组OCT的反应相同。稀释试验显示稀释倍数与浓度之间具有良好的线性关系。添加的不同浓度的OCT平均回收率为90.4%。ELISA中OCT蛋白水平与OCT酶活性之间存在良好的相关性（r = 0.987，p < 0.0001）。慢性肝炎患者（110.7±80 ng/ml）与健康受试者（34.4±20.7 ng/ml）之间观察到OCT血清水平存在显著差异（p < 0.0001）。健康受试者中OCT血清水平在性别之间存在显著差异（p < 0.0001）。