Mitchell R, Lambert A, Crosby S R, White A, Robertson W R
Department of Clinical Biochemistry (Medicine), University of Manchester, Hope Hospital, Salford, UK.
Ann Clin Biochem. 1990 Jan;27 ( Pt 1):59-64. doi: 10.1177/000456329002700112.
We have previously reported a bioassay for human plasma ACTH based upon trypsin dispersed guinea-pig adrenal cells which was sensitive to 100 ng/L ACTH in unextracted human plasma when measured against human pituitary ACTH (1-39) standard 74/555. We now present a bioassay of increased sensitivity (12 ng/L) which incorporates three major changes. The trypsin/trypsin inhibitor step in the cell dispersion protocol has been replaced with collagenase, donor calf serum (3%) has been incorporated into the standard curve and ACTH has been extracted from human plasma and dilutions of standard hormone by a sephacryl bound monoclonal antibody (2A3) directed towards the 25-39 sequence. The extracted standard curve has a detection limit of 6 ng/L and the cells can tolerate up to 50% plasma equivalent concentration. Thus, the improved assay has a detection limit of 12 ng/L ACTH in plasma. The assay can now measure bioactive plasma ACTH levels reliably in the normal range.
我们之前报道过一种基于胰蛋白酶分散豚鼠肾上腺细胞的人血浆促肾上腺皮质激素（ACTH）生物测定法，当以人垂体ACTH（1-39）标准品74/555为对照时，该方法对未提取的人血浆中100 ng/L的ACTH敏感。我们现在提出一种灵敏度更高（12 ng/L）的生物测定法，该方法有三个主要变化。细胞分散方案中的胰蛋白酶/胰蛋白酶抑制剂步骤已被胶原酶取代，标准曲线中加入了供体小牛血清（3%），并且已通过针对25-39序列的琼脂糖凝胶结合单克隆抗体（2A3）从人血浆和标准激素稀释液中提取ACTH。提取的标准曲线检测限为6 ng/L，细胞可耐受高达50%血浆等效浓度。因此，改进后的测定法血浆中ACTH的检测限为12 ng/L。该测定法现在可以在正常范围内可靠地测量生物活性血浆ACTH水平。
