大鼠生长激素的酶免疫测定法：验证及其在血浆水平测定和体外释放测定中的应用

An enzyme immunoassay for rat growth hormone: validation and application to the determination of plasma levels and in vitro release.

作者信息

Ezan E, Laplante E, Bluet-Pajot M T, Mounier F, Mamas S, Grouselle D, Grognet J M, Kordon C

机构信息

CEA, Service de Pharmacologie et d'Immunologie, Gif-sur-Yvette, France.

出版信息

J Immunoassay. 1997 Nov;18(4):335-56. doi: 10.1080/01971529708005826.

DOI:10.1080/01971529708005826

PMID:9358340

Abstract

A competitive enzyme immunoassay for rat growth hormone (rGH) has been developed using polyclonal anti-rGH antibodies and an acetylcholinesterase (EC 3.1.1.7.) enzymatic tracer coupled covalently with rGH. The assay was performed in 96-well microtiter plates coated with rabbit polyclonal anti-goat immunoglobulin antibodies. Molecular sieve filtration and Western blot analysis revealed a single immunoreactive peak for rat plasma or pituitary extracts. Cross-reactivity with other rat pituitary hormones or human GH was less than 1%. Assay of samples in a concentration range of 0.7 to 69 ng/ml by enzyme immunoassay and radioimmunoassay were well correlated (r = 0.87 and 0.85 respectively for plasma and culture medium samples). Intra- and inter-assay variations in plasma were 4 (n = 24) and 14% (n = 9) respectively. Minimal detectable amounts of rGH were 0.6 ng/ml. A two-site immunometric assay also developed with the same antibodies allowed a detection threshold of 0.25 ng/ml.

摘要

利用多克隆抗大鼠生长激素（rGH）抗体和与rGH共价偶联的乙酰胆碱酯酶（EC 3.1.1.7.）酶标记物，开发了一种用于大鼠生长激素（rGH）的竞争性酶免疫测定法。该测定在涂有兔多克隆抗山羊免疫球蛋白抗体的96孔微量滴定板中进行。分子筛过滤和蛋白质印迹分析显示大鼠血浆或垂体提取物有一个单一的免疫反应峰。与其他大鼠垂体激素或人GH的交叉反应率小于1%。通过酶免疫测定法和放射免疫测定法对浓度范围为0.7至69 ng/ml的样品进行测定，两者相关性良好（血浆和培养基样品的r分别为0.87和0.85）。血浆中的批内和批间变异分别为4%（n = 24）和14%（n = 9）。rGH的最低检测量为0.6 ng/ml。用相同抗体开发的双位点免疫测定法的检测阈值为0.25 ng/ml。