QUANTIFICATION OF ANTIGEN-SPECIFIC CD8(+) T-CELLS USING AN ELISPOT ASSAY

被引：315

作者：

MIYAHIRA, Y ^{[1
]}

MURATA, K ^{[1
]}

RODRIGUEZ, D ^{[1
]}

RODRIGUEZ, JR ^{[1
]}

ESTEBAN, M ^{[1
]}

RODRIGUES, MM ^{[1
]}

ZAVALA, F ^{[1
]}

机构：

[1] CSIC,CTR NACL BIOTECNOL,E-28049 MADRID,SPAIN

来源：

JOURNAL OF IMMUNOLOGICAL METHODS | 1995年 / 181卷 / 01期

基金：

美国国家卫生研究院;

关键词：

CD8(+) T CELL; ELISPOT; IFN-GAMMA; SYNTHETIC PEPTIDE; RECOMBINANT VACCINIA VIRUS; MALARIA;

D O I：

10.1016/0022-1759(94)00327-S

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

An ELISPOT assay to detect and determine the number of antigen specific CD8(+) T cells was standardized using cloned murine CD8(+) T cells specific for the epitope SWPSAEQI of a rodent malaria antigen. This assay is based on the detection of IFN-gamma secretion by single cells after their stimulation with antigen. The interferon secretion is visualized as spots revealed by using enzyme labeled anti-IFN-gamma monoclonal antibodies. Using known numbers of cloned murine CD8(+) T cells it was determined that the assay detects 80-95% of these CD8(+) T cells. The optimal culture conditions for the stimulation of the CD8(+) T cells were determined and the antigen concentration, number of antigen presenting cells and supplement of growth factors required to perform the assay were defined. This ELISPOT assay can be performed with spleen cells from immunized mice, and provide the precise number of antigen specific CD8(+) T cells present in mixed lymphocyte populations. This method is more sensitive than the chromium-51 release assay, and much simpler than the conventional precursor frequency analysis, providing the number of antigen specific CD8(+) T cells in 36-48 h.

引用

页码：45 / 54

页数：10

共 12 条

[1] PHENOTYPIC ANALYSIS OF THE INFLAMMATORY EXUDATE IN MURINE LYMPHOCYTIC CHORIOMENINGITIS [J].