磁分离酶联免疫荧光法(MEIF)检测人血清中胰岛素

Establishment of the magnetic separation enzyme immunofluorescence method for detecting insulin

目的：采用磁微粒分离酶联免疫荧光（MEIF）分析技术建立一种简便、高敏感性、定量检测人血清胰岛素（insu-lin）的新方法。方法：选用2株识别人胰岛素不同表位的单克隆抗体（mAb）。一株mAb用异硫氰酸荧光素（FITC）标记,另一株用碱性磷酸酶（AP）标记;偶联羊抗FITC抗体的磁珠用作固相分离载体,4-甲基磷酸伞型酮用作荧光底物。结果：成功建立了定量检测人血清胰岛素的MEIF,灵敏度2·0μIu/mL,线形范围0-188·52μIu/mL,批内变异系数（CV）为4·3%-5·2%,批间CV为2·6%-9·5%,稀释回收率为93%-117%,加标回收率为94%-113%。实际样品的测定结果与倍爱康公司商品化人insulin磁分离发光系统检测试剂盒的检测结果比较,具有良好的相关性。结论：MEIF定量测定人insulin方法成本低、灵敏度高、稳定性好,在临床免疫检测领域具有广阔的应用前景。

AIM： To establish a convenient and sensitive magnetic separation enzyme immunofluorescence （MEIF） method for detecting human insulin. METHODS： Two monoclonal antibodies （mAbs） were conjugated with FITC and alkaline phosphatase （AP） respectively, which were incorporated magnetic solid phase separation. Magnetic beads were coupled with sheep anti-FITC antibody as solid phase and 4-Methylumbelliferyl-phosphoric acid （4- MUP） was used as substrate to set up MEIF for detecting insulin. RESULTS： The sensitivity of this assay was 2.0 μlu/mL, the linear range was from 0 μlu/mL to 188.52 μlu/ mL, and the intra-assay variation and inter-assay variation were 4.3% - 5.2% and 2.6% - 9.5%, respectively. The recovery rate of dilution was 92.6% - 117% and the recovery rate of accession was 106% - 121%. The result of the assay correlated well with that of magnetic enzyme chemiluminescence immunoassay system. CONCLUSION： The MEIF for detecting insulin is low at cost, sensitive, specific and stable, which can be widely used in clinical immune detection.