建立一种使用荧光底物的白细胞介素-1α高灵敏度酶联免疫吸附测定法。

Establishment of a highly sensitive enzyme-linked immunosorbent assay for interleukin-1 alpha employing a fluorogenic substrate.

作者信息

Mukaida N, Kasahara T, Ko Y, Kawai T

机构信息

Department of Clinical Pathology, Jichi Medical School, Tochigi-ken, Japan.

出版信息

J Immunol Methods. 1988 Feb 24;107(1):41-6. doi: 10.1016/0022-1759(88)90006-3.

DOI:10.1016/0022-1759(88)90006-3

PMID:3257779

Abstract

We have previously established a non-competitive solid-phase enzyme-linked immunosorbent assay (ELISA) specific for interleukin-1 alpha (IL-1 alpha) using a combination of polyclonal antibody as the immobilized antibody, biotinylated monoclonal antibody as the second antibody and avidin-peroxidase. The level of detection of that ELISA was 200-500 pg/ml. In order to improve its sensitivity, we have used streptavidin-beta-D-galactosidase and the fluorogenic substance 4-methylumbelliferyl-D-galactopyranoside as enzyme substrate. With this system IL-1 alpha could be detected at concentrations as low as 10-50 pg/ml, which was about 10-20 times more sensitive than conventional mouse thymocyte co-stimulator assays. Furthermore, the assay system was specific for IL-1 alpha in that neither IL-1 beta nor interleukin-2 (IL-2) interfered.

摘要

我们之前建立了一种非竞争性固相酶联免疫吸附测定法（ELISA），该方法专门用于检测白细胞介素-1α（IL-1α），使用多克隆抗体作为固定抗体，生物素化单克隆抗体作为二抗，以及抗生物素蛋白-过氧化物酶。该ELISA的检测水平为200 - 500 pg/ml。为了提高其灵敏度，我们使用了链霉亲和素-β-D-半乳糖苷酶和荧光物质4-甲基伞形基-D-吡喃半乳糖苷作为酶底物。使用该系统，可检测到低至10 - 50 pg/ml浓度的IL-1α，其灵敏度比传统的小鼠胸腺细胞共刺激测定法高约10 - 20倍。此外，该测定系统对IL-1α具有特异性，因为白细胞介素-1β（IL-1β）和白细胞介素-2（IL-2）均不产生干扰。