Open AccessSimultaneous cytometric analysis for the expression of cytoplasmic and surface antigens in activated T cells
Author(s) -
Hayden Georgina E.,
Walker Karen Z.,
Wotherspoon John S.,
Raison Robert L.,
Miller Jacques F. A. P.
Publication year - 1988
Publication title -
cytometry
Language(s) - English
Resource type - Journals
eISSN - 1097-0320
pISSN - 0196-4763
DOI - 10.1002/cyto.990090108
Subject(s) - cytoplasm , antigen , expression (computer science) , flow cytometry , microbiology and biotechnology , biology , chemistry , computer science , immunology , programming language
A method of two‐colour immunofluorescence staining has been developed to allow the simultaneous analysis of both surface and cytoplasmic antigens. This involves the use of direct fluorochrome antibody conjugates for cell‐surface antigen staining, followed by cell permeabilization and the staining of cytoplasmic antigens with biotinylated antibodies and streptavidin‐fluorochrome conjugates. Fluorochrome‐antibody conjugates bound to cell‐surface epitopes were found not to be affected by the subsequent permeabilisation and cytoplasmic staining. This method was used to examine the surface phenotype of T cells expressing a cytoplasmic antigen, STA. STA is a unique determinant detected in activated human T cells by the monoclonal antibody K‐1‐21, which also recognizes a cross‐reactive conformation‐dependent epitope on human free kappa light chains. Cytometric analysis showed that STA is found in both Leu 2a + cytotoxic/suppressor T cells and Leu 3a + helper/inducer T cells but is not induced in the Leu 15 + population which contains suppressor T cells. STA was also shown to be an activation antigen in murine T cells.