使用表达β-半乳糖苷酶的K562靶细胞系进行人自然杀伤细胞活性的酶释放测定。

Enzyme release assay of human NK cell activity using beta-galactosidase-expressing K562 target cell line.

作者信息

Ohmori H, Ikeda H, Tanigawa T, Takai T, Hikida M

机构信息

Department of Biotechnology, Faculty of Engineering, Okayama University, Japan.

出版信息

J Immunol Methods. 1993 Aug 26;164(1):131-5. doi: 10.1016/0022-1759(93)90283-d.

DOI:10.1016/0022-1759(93)90283-d

PMID:8360503

Abstract

In the present report, we established a K562 cell line useful for an enzyme release assay of human natural killer (NK) activity. Human myelogenous leukemia cell line, K562, was transfected with a plasmid carrying Escherichia coli beta-galactosidase (beta-gal) gene. A colony that permanently expresses the enzyme activity was isolated, and designated K562/Zneo. Incubation of K562/Zneo cells (1 x 10(4)) with nonadherent human peripheral blood lymphocytes (PBL) resulted in the release of beta-gal activity depending on the incubation time and the number of effector cells. Released beta-gal activity was assayed sensitively by using 4-methylumbelliferyl-beta-D-galactoside, a fluorescent substrate. The cytolytic activity of PBL was augmented significantly when the cells were preincubated with interleukin-2 for 20 h. This enzyme release assay showed a comparable sensitivity to that of 51Cr release assay. Thus, K562/Zneo cell line is thought to be useful for the nonradioactive assay of human NK and lymphokine-activated killer activities.

摘要

在本报告中，我们建立了一种可用于人自然杀伤（NK）活性酶释放测定的K562细胞系。用人髓性白血病细胞系K562转染携带大肠杆菌β-半乳糖苷酶（β-gal）基因的质粒。分离出一个永久表达酶活性的克隆，并命名为K562/Zneo。将K562/Zneo细胞（1×10⁴）与非贴壁人外周血淋巴细胞（PBL）一起孵育，会导致β-gal活性的释放，其依赖于孵育时间和效应细胞的数量。使用荧光底物4-甲基伞形酮基-β-D-半乳糖苷对释放的β-gal活性进行灵敏测定。当细胞与白细胞介素-2预孵育20小时时，PBL的细胞溶解活性显著增强。这种酶释放测定显示出与⁵¹Cr释放测定相当的灵敏度。因此，K562/Zneo细胞系被认为可用于人NK和淋巴因子激活的杀伤活性的非放射性测定。