Modified ELISPOT technique--highly significant inverse correlation of post-Tx donor-reactive IFNgamma-producing cell frequencies with 6 and 12 months graft function in kidney transplant recipients.
BACKGROUND
Upcoming trials for immunosuppression minimization and tolerance induction require the development of reliable in vitro assays for monitoring cellular alloimmunity in transplant patients. The IFN-gamma ELISPOT assay represents a promising tool for monitoring alloreactive memory/effector T cells. As T lymphopenia is a common finding during the early post-transplant (post-Tx) period, the IFN-gamma ELISPOT technique was here modified by using ELISPOT responder cells with enhanced percentage and standardized number of 200,000 T cells per well.


METHODS
Peripheral blood mononuclear cells (PBMNC) of kidney transplant recipients were depleted of CD14+ and CD15+ cells to increase the percentage of T cells from average 47.8% to 71.5% before transplantation (pre-Tx) and from 39.7% to 74.9% post-Tx. The assay was tested in a population of 23 de novo renal transplant patients for clinical relevance. Before and at 2-3 times during the first 6 months post-Tx, IFN-gamma-producing donor-reactive as well as recall antigen-reactive cell frequencies (Candida, tuberculin, tetanus) were determined and correlated with outcome.


RESULTS
Pre-Tx donor-reactive ELISPOT frequencies were enhanced in patients with acute rejection compared to non-rejectors. Moreover, mean post-Tx donor-reactive ELISPOT frequencies showed a highly significant inverse correlation with renal function at 6 and 12 months. In contrast, recall antigen-reactive ELISPOT frequencies did not correlate with outcome.


CONCLUSIONS
Our results suggest that the modified donor-reactive ELISPOT approach might provide a useful surrogate marker for renal transplant outcome with possible utility especially in T-lymphopenic patients.
Näther
Bele Johanna
BJ
Institute of Medical Immunology, Charité Campus Mitte, Schumannstrasse 20/21, 10098 Berlin, Germany.
Nickel
Peter
P
Bold
Gantuja
G
Presber
Franziska
F
Schönemann
Constanze
C
Pratschke
Johann
J
Volk
Hans-Dieter
HD
Reinke
Petra
P
D015415:Biomarkers / Q000032:analysis; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006084:Graft Rejection / Q000276:immunology*; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism*; D016030:Kidney Transplantation / Q000276:immunology*; D013601:T-Lymphocytes / Q000166:cytology* / Q000276:immunology*
D015415:Biomarkers; D007371:Interferon-gamma