Howell S, Murray H, Scott C S, Turner A J, Kenny A J
Department of Biochemistry, University of Leeds, UK.
Biochem J. 1991 Sep 1;278 ( Pt 2)(Pt 2):417-21. doi: 10.1042/bj2780417.
Endopeptidase-24.11 is a widely distributed cell-surface enzyme with a key role in the metabolism of neuropeptides. It is now known to be identical with CD-10, the common acute-lymphoblastic-leukaemia antigen (CALLA). An e.l.i.s.a. is described which utilizes two antibodies, one monoclonal, the other polyclonal, generated to pig endopeptidase-24.11. These antibodies cross-reacted with human endopeptidase-24.11, thus making the assay applicable to both species. By using optimum conditions for the e.l.i.s.a., as little as 25 pg of pure pig endopeptidase-24.11 could be quantified at 95% confidence limits. E.l.i.s.a. of tissue homogenates from a variety of pig tissues and of human kidney correlated well with enzymic assays. However, the use of detergents to solubilize the antigen greatly decreased the sensitivity of the e.l.i.s.a. The e.l.i.s.a. is 1000-fold more sensitive than the immunoradiometric assay and has advantages in specificity over enzymic assays. Daudi cells, some leukaemic cells shown to be CALLA-positive, and Caco-2 cells, could also be assayed, but N2 cavitation was necessary to fragment the cells, and only part of the total endopeptidase-24.11 activity in Daudi cells was recognized by the e.l.i.s.a.
内肽酶-24.11是一种广泛分布的细胞表面酶,在神经肽代谢中起关键作用。现已知道它与普通急性淋巴细胞白血病抗原(CALLA)即CD-10相同。本文描述了一种酶联免疫吸附测定法(ELISA),该方法利用针对猪内肽酶-24.11产生的两种抗体,一种是单克隆抗体,另一种是多克隆抗体。这些抗体与人类内肽酶-24.11发生交叉反应,因此该测定法适用于这两个物种。通过使用ELISA的最佳条件,在95%置信限下,低至25皮克的纯猪内肽酶-24.11都可以被定量。对来自各种猪组织的组织匀浆和人肾进行ELISA测定,结果与酶活性测定结果高度相关。然而,使用去污剂溶解抗原会大大降低ELISA的灵敏度。该ELISA比免疫放射测定法灵敏1000倍,并且在特异性方面优于酶活性测定法。也可以对Daudi细胞(一些显示为CALLA阳性的白血病细胞)和Caco-2细胞进行测定,但需要进行氮空化以使细胞破碎,并且ELISA只能识别Daudi细胞中总内肽酶-24.11活性的一部分。