Flow-cytometric detection of vaccinia-induced memory effector CD4(+), CD8(+), and gamma delta TCR(+) T cells capable of antigen-specific expansion and effector functions.
We developed a carboxyfluorescein succinimidyl ester (CFSE)-based flow-cytometric assay that can detect different subsets of vaccinia-specific T cells capable of both antigen-specific expansion and protective effector functions. Proliferation and effector functions were detected by CFSE dilution and intracellular staining, respectively. Absolute numbers of CD4(+)/CFSE(lo)/interferon (IFN)- gamma (+), CD8(+)/CFSE(lo)/IFN- gamma (+), CD8(+)/CFSE(lo)/granzyme A(+), and CD8(+)/CFSE(lo)/CD107a(+) T cells present after in vitro stimulation with live vaccinia were significantly higher in immunized individuals (P<.05). These CD4(+) and CD8(+) T cell increases were >2 log higher than increases detectable by standard lymphoproliferation and cytotoxicity assays. Vaccinia-specific CD8(+)/CFSE(lo)/IFN- gamma (+) and granzyme A(+) T cell responses were significantly correlated with the results of standard (51)Cr-release cytolytic assays (P<.05). Furthermore, vaccinia induced antigen-specific memory gamma delta T cells. We demonstrate that vaccinia induces robust memory effector CD4(+), CD8(+), and gamma delta T cells, all of which are relevant for protection against smallpox. CFSE-based flow-cytometric assays will be useful in evaluating cell-mediated immune responses induced by new smallpox vaccines.
Abate
Getahun
G
Department of Internal Medicine, Saint Louis University, St. Louis, Missouri 63110, USA.
Eslick
Joy
J
Newman
Frances K
FK
Frey
Sharon E
SE
Belshe
Robert B
RB
Monath
Thomas P
TP
Hoft
Daniel F
DF
D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D005434:Flow Cytometry; D005452:Fluoresceins / Q000378:metabolism; D006801:Humans; D007156:Immunologic Memory; D016692:Receptors, Antigen, T-Cell, gamma-delta / Q000276:immunology*; D013388:Succinimides / Q000378:metabolism; D014615:Vaccinia / Q000276:immunology*; D014616:Vaccinia virus / Q000235:genetics / Q000276:immunology
C087165:5-(6)-carboxyfluorescein diacetate succinimidyl ester; D005452:Fluoresceins; D016692:Receptors, Antigen, T-Cell, gamma-delta; D013388:Succinimides