TY  - JOUR
A1  - Wunsch, Marie
A1  - Zhang, Wenji
A1  - Hanson, Jodi
A1  - Caspell, Richard
A1  - Karulin, Alexey Y.
A1  - Recks, Mascha S.
A1  - Kuerten, Stefanie
A1  - Sundararaman, Srividya
A1  - Lehmann, Paul V.
T1  - Characterization of the HCMV-Specific CD4 T Cell Responses that Are Associated with Protective Immunity
JF  - Viruses
N2  - Most humans become infected with human cytomegalovirus (HCMV). Typically, the immune system controls the infection, but the virus persists and can reactivate in states of immunodeficiency. While substantial information is available on the contribution of CD8 T cells and antibodies to anti-HCMV immunity, studies of the T\(_{H}\)1, T\(_{H}\)2, and T\(_{H}\)17 subsets have been limited by the low frequency of HCMV-specific CD4 T cells in peripheral blood mononuclear cell (PBMC). Using the enzyme-linked Immunospot\(^{®}\) assay (ELISPOT) that excels in low frequency measurements, we have established these in a sizable cohort of healthy HCMV controllers. Cytokine recall responses were seen in all seropositive donors. Specifically, interferon (IFN)-\({\gamma}\) and/or interleukin (IL)-17 were seen in isolation or with IL-4 in all test subjects. IL-4 recall did not occur in isolation. While the ratios of T\(_{H}\)1, T\(_{H}\)2, and T\(_{H}\)17 cells exhibited substantial variations between different individuals these ratios and the frequencies were relatively stable when tested in samples drawn up to five years apart. IFN-\({\gamma}\) and IL-2 co-expressing polyfunctional cells were seen in most subjects. Around half of the HCMV-specific CD4 cells were in a reversible state of exhaustion. The data provided here established the T\(_{H}\)1, T\(_{H}\)2, and T\(_{H}\)17 characteristic of the CD4 cells that convey immune protection for successful immune surveillance against which reactivity can be compared when the immune surveillance of HCMV fails.
KW  - memory cells
KW  - hcv infection
KW  - signature
KW  - Enzyme-Linked Immunospot assay (ELISPOT)
KW  - cytokine secretion kinetics
KW  - chronic viral infection
KW  - HCMV infection
KW  - CD4 T cells
KW  - exhaustion
KW  - activation
KW  - human cytomegalovirus (HCMV)
KW  - B cells
KW  - cytomegalovirus
KW  - elispot
Y1  - 2015
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-151462
VL  - 7
SP  - 4414
EP  - 4437
ER  - 
TY  - JOUR
A1  - Karulin, Alexey Y.
A1  - Karacsony, Kinga
A1  - Zhang, Wenji
A1  - Targoni, Oleg S.
A1  - Moldova, Ioana
A1  - Dittrich, Marcus
A1  - Sundararaman, Srividya
A1  - Lehmann, Paul V.
T1  - ELISPOTs produced by CD8 and CD4 cells follow Log Normal size distribution permitting objective counting
JF  - Cells
N2  - Each positive well in ELISPOT assays contains spots of variable sizes that can range from tens of micrometers up to a millimeter in diameter. Therefore, when it comes to counting these spots the decision on setting the lower and the upper spot size thresholds to discriminate between non-specific background noise, spots produced by individual T cells, and spots formed by T cell clusters is critical. If the spot sizes follow a known statistical distribution, precise predictions on minimal and maximal spot sizes, belonging to a given T cell population, can be made. We studied the size distributional properties of IFN-γ, IL-2, IL-4, IL-5 and IL-17 spots elicited in ELISPOT assays with PBMC from 172 healthy donors, upon stimulation with 32 individual viral peptides representing defined HLA Class I-restricted epitopes for CD8 cells, and with protein antigens of CMV and EBV activating CD4 cells. A total of 334 CD8 and 80 CD4 positive T cell responses were analyzed. In 99.7% of the test cases, spot size distributions followed Log Normal function. These data formally demonstrate that it is possible to establish objective, statistically validated parameters for counting T cell ELISPOTs.
KW  - ELISPOT
KW  - software
KW  - IFN-γ
KW  - IL-17
KW  - T cells
KW  - Normal Distribution
KW  - spot size
KW  - gating
KW  - cytokines
KW  - IL-2
KW  - IL-4
KW  - IL-5
KW  - CD8
KW  - CD4
Y1  - 2015
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-149648
VL  - 4
IS  - 1
ER  -