一种可快速灵敏地通过光度法测定单核吞噬细胞对红细胞吞噬作用的方法。

A rapid and sensitive method allowing photometric determination of erythrophagocytosis by mononuclear phagocytes.

作者信息

Jungi T W

出版信息

J Immunol Methods. 1985 Sep 3;82(1):141-53. doi: 10.1016/0022-1759(85)90233-9.

Abstract

A spectrometric assay for assessing erythrophagocytosis by mononuclear phagocytes is described. It is based on the haemoglobin-catalyzed conversion of a benzidine derivative into a coloured product in the presence of H2O2 (pseudoperoxidase activity). The assay is set up in microtitre plates, and following an uptake phase and removal of non-ingested erythrocytes, pseudoperoxidase activity is measured in detergent lysates of phagocytes, using an ELISA reader photometer. Various detergents and substrates were evaluated. SDS was found to be the most suitable detergent. Diaminobenzidine (in PBS, pH 7.4) was the substrate of choice for enumerating ingested erythrocytes in a range from 10(4) to 5-8X10(5) sheep erythrocytes. Ortho-tolidine (in acetate buffer, pH 5.5) could be used in a range from 2X10(3) to 2X10(5) sheep erythrocytes. The results obtained with human peripheral blood monocytes or monocyte-derived macrophages and IgG-sensitized sheep erythrocytes correlated well with those obtained using 51Cr-labelled, IgG-sensitized erythrocytes.

摘要

描述了一种用于评估单核吞噬细胞红细胞吞噬作用的光谱分析方法。它基于在过氧化氢存在下血红蛋白催化联苯胺衍生物转化为有色产物（假过氧化物酶活性）。该分析在微量滴定板中进行，经过摄取阶段并去除未摄取的红细胞后，使用酶联免疫吸附测定仪光度计在吞噬细胞的去污剂裂解物中测量假过氧化物酶活性。评估了各种去污剂和底物。发现十二烷基硫酸钠是最合适的去污剂。二氨基联苯胺（在pH 7.4的磷酸盐缓冲盐水中）是用于计数10⁴至5 - 8×10⁵个绵羊红细胞范围内摄取红细胞的首选底物。邻联甲苯胺（在pH 5.5的醋酸盐缓冲液中）可用于2×10³至2×10⁵个绵羊红细胞的范围。用人外周血单核细胞或单核细胞衍生的巨噬细胞以及IgG致敏的绵羊红细胞获得的结果与使用⁵¹Cr标记的、IgG致敏的红细胞获得的结果相关性良好。