MEASUREMENT OF TUMOR-NECROSIS-FACTOR ACTIVITY BY FLOW-CYTOMETRY

被引：20

作者：

LEVESQUE, A ^{[1
]}

PAQUET, A ^{[1
]}

PAGE, M ^{[1
]}

机构：

[1] UNIV LAVAL,FAC MED,DEPT BIOCHEM,ST FOY,PQ G1K 7P4,CANADA

来源：

CYTOMETRY | 1995年 / 20卷 / 02期

关键词：

BIOASSAY; CYTOTOXICITY; ETHIDIUM HOMODIMER-1; CALCEIN-AM; CALCEIN;

D O I：

10.1002/cyto.990200211

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

Tumor necrosis factor-alpha (TNF-alpha) is a monokine of 17 kDa produced by activated macrophages and various cells involved in the immune system, We propose a new method for the measurement of TNF activity using flow cytometry. After an incubation with TNF, L929 cells were harvested and treated with a calcein-AM and ethidium homodimer-1 solution, Nonfluorescent calcein-AM is hydrolyzed by intracellular esterases to yield fluorescent calcein. The ethidium homodimer-1 is a high-affinity red fluorescent DNA dye that is internalized only through altered cell membranes. A very good correlation was observed between the calcein fluorescence in-tensity and the number of viable cells as well as the ethidium fluorescence and the number of cells with altered membranes. The assay is sensitive, inexpensive, and correlates with the already reported crystal violet assay while measuring membrane alteration by TNF, It allows the simultaneous measurement of total living and dead cells. There is no interference with culture medium components, This method is rapid and may be used for routine measurement of TNF activity. (C) 1995 Wiley-Liss, Inc.

引用

页码：181 / 184

页数：4

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