Detection of low-frequency antigen-specific IL-10-producing CD4(+) T cells via ELISPOT in PBMC: cognate vs. nonspecific production of the cytokine.
Single-cell resolution cytokine ELISPOT assays are increasingly used to gain insights into clonal sizes of type 1 and type 2 effector T cell populations in vivo. However, ELISPOT assays permitting monitoring of regulatory IL-10-producing T cells have so far not been established. Unlike IFN-gamma, IL-2, IL-4, and IL-5 assays performed on PBMC in which the recall antigen-induced cytokine spots are T cell-derived, we show here that in such assays IL-10 is primarily monocyte-derived. T cell-derived IL-10 spots were 80 x 10(3) microm(2) in size, seven times larger than spots produced by monocytes, and B cells produced even smaller spots. Based on spot size gating and the use of B cells as APC, we have established test conditions that permit measurement of cognate IL-10 production by low-frequency antigen-specific T cells. IL-10-producing PPD-specific CD4(+) T cells were detected in frequencies comparable to IFN-gamma-secreting CD4(+) T cells in tuberculosis patients, but not in uninfected healthy control individuals. In contrast, IL-10-secreting CD4(+) T cells specific for a panel of recall antigens could not be detected in frequencies >1/100,000 in healthy individuals whose CD4(+) cells responded to these antigens with type 1 or type 2 cytokine production in the 1:100,000-1:1000 frequency range. Therefore, the induction of IL-10-producing T cells seems to be under tighter control than that of Th1/Th2 cells, apparently confined to states of chronic immune stimulation. Access to low-frequency immune monitoring of IL-10-producing T cells will provide new insights into the role of regulatory T cells in health and disease.
Guerkov
Robert E M
RE
Department of Pathology, Case Western Reserve University School of Medicine, Cleveland, OH 44106-4943, USA.
Targoni
Oleg S
OS
Kreher
Christian R
CR
Boehm
Bernhard O
BO
Herrera
Maria T
MT
Tary-Lehmann
Magdalena
M
Lehmann
Paul V
PV
Schwander
Stephan K
SK
D015496:CD4-Positive T-Lymphocytes / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D016753:Interleukin-10 / Q000378:metabolism*; D016176:T-Lymphocyte Subsets / Q000378:metabolism*
D016753:Interleukin-10