Ida N, Sakurai S, Kawano G
Medical Devices and Diagnostics Research Laboratories, Toray Industries Inc., Kanagawa, Japan.
Cytokine. 1994 Jan;6(1):32-9. doi: 10.1016/1043-4666(94)90005-1.
We developed a highly sensitive enzyme-linked immunosorbent assay (ELISA) for human monocyte chemotactic and activating factor (MCAF), an inflammatory cytokine that plays an important role in the recruitment of blood monocytes to areas of inflammation. The ELISA, which is based on a sandwich method using two newly-developed monoclonal antibodies, could quantitatively detect MCAF in the range between 2.5 pg/ml (50 fg/sample) to 300 pg/ml after incubation for a total of 2 h, and showed no cross-reactivity with various structurally-related IL-8 superfamily proteins. It was not affected by blood or urine components non-specifically, and thus was directly applicable to clinical specimens. When serum and urine samples from healthy subjects were measured, they all turned out to contain detectable levels of MCAF (more than 30 pg/ml). By gel-filtration column chromatography analysis, MCAF in the body fluids was eluted as a single peak at the position corresponding to the molecular weight of 10 kD, suggesting that it exists as a monomer form, free from carrier proteins. The established ELISA here is expected to be effectively used for the further investigations on the relationship of MCAF with various inflammatory diseases.
我们开发了一种用于检测人单核细胞趋化激活因子（MCAF）的高灵敏度酶联免疫吸附测定（ELISA）方法。MCAF是一种炎症细胞因子，在将血液单核细胞募集到炎症部位的过程中发挥重要作用。该ELISA方法基于使用两种新开发的单克隆抗体的夹心方法，在总共孵育2小时后，能够定量检测2.5 pg/ml（50 fg/样本）至300 pg/ml范围内的MCAF，并且与各种结构相关的IL-8超家族蛋白无交叉反应。它不受血液或尿液成分的非特异性影响，因此可直接应用于临床标本。对健康受试者的血清和尿液样本进行检测时，结果显示它们均含有可检测水平的MCAF（超过30 pg/ml）。通过凝胶过滤柱色谱分析，体液中的MCAF在对应于10 kD分子量的位置以单峰形式洗脱，表明它以单体形式存在，不与载体蛋白结合。这里建立的ELISA方法有望有效地用于进一步研究MCAF与各种炎症性疾病的关系。
