PMID- 25149304
OWN - NLM
STAT- MEDLINE
DCOM- 20150511
LR  - 20181202
IS  - 1940-6029 (Electronic)
IS  - 1064-3745 (Linking)
VI  - 1186
DP  - 2014
TI  - CTL ELISPOT assay.
PG  - 75-86
LID - 10.1007/978-1-4939-1158-5_6 [doi]
AB  - Enzyme-linked immune absorbent spot (Elispot) is a quantitative method for 
      measuring relevant parameters of T cell activation. The sensitivity of Elispot 
      allows the detection of low-frequency antigen-specific T cells that secrete 
      cytokines and effector molecules, such as granzyme B and perforin. Cytotoxic T 
      cell (CTL) studies have taken advantage with this high-throughput technology by 
      providing insights into quantity and immune kinetics. Accuracy, sensitivity, 
      reproducibility, and robustness of Elispot resulted in a wide range of 
      applications in research as well as in diagnostic field. Actually, CTL monitoring 
      by Elispot is a gold standard for the evaluation of antigen-specific T cell 
      immunity in clinical trials and vaccine candidates where the ability to detect 
      rare antigen-specific T cells is of relevance for immune diagnostic. The most 
      utilized Elispot assay is the interferon-gamma (IFN-gamma) test, a marker for CD8(+) 
      CTL activation, but Elispot can also be used to distinguish different subsets of 
      activated T cells by using other cytokines such as T-helper (Th) 1-type cells 
      (characterized by the production of IFN-gamma, IL-2, IL-6, IL-12, IL-21, and TNF-alpha), 
      Th2 (producing cytokines like IL-4, IL-5, IL-10, and IL-13), and Th17 (IL-17) 
      cells. The reliability of Elispot-generated data, by the evaluation of T cell 
      frequency recognizing individual antigen/peptide, is the core of this method 
      currently applied widely to investigate specific immune responses in cancer, 
      infections, allergies, and autoimmune diseases. The Elispot assay is competing 
      with other methods measuring single-cell cytokine production, e.g., intracellular 
      cytokine by FACS or Miltenyi cytokine secretion assay. Other types of lymphocyte 
      frequency and function assays include limiting dilution assay (LDA), cytotoxic T 
      cell assay (CTL), and tetramer staining. Compared with respect to sensitivity the 
      Elispot assay is outranking other methods to define frequency of antigen-specific 
      lymphocytes. The method described herein would like to offer helpful and clear 
      protocols for researchers that apply Elispot. IFN-gamma and perforin Elispot assays 
      are described.
FAU - Ranieri, Elena
AU  - Ranieri E
AD  - Department of Surgical and Medical Sciences, School of Medicine, University of 
      Foggia, Ospedali Riuniti, Viale Luigi Pinto, 1, Foggia, 71122, Italy, 
      elena.ranieri@unifg.it.
FAU - Popescu, Iulia
AU  - Popescu I
FAU - Gigante, Margherita
AU  - Gigante M
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - United States
TA  - Methods Mol Biol
JT  - Methods in molecular biology (Clifton, N.J.)
JID - 9214969
RN  - 0 (Antibodies, Monoclonal)
RN  - 0 (Epitopes, T-Lymphocyte)
RN  - 126465-35-8 (Perforin)
RN  - 82115-62-6 (Interferon-gamma)
RN  - EC 3.4.21.- (Granzymes)
SB  - IM
MH  - Antibodies, Monoclonal/immunology
MH  - Cytomegalovirus/immunology
MH  - Enzyme-Linked Immunospot Assay/*methods
MH  - Epitopes, T-Lymphocyte/immunology
MH  - Granzymes/metabolism
MH  - Herpesvirus 4, Human/immunology
MH  - High-Throughput Screening Assays/methods
MH  - Humans
MH  - Interferon-gamma/*analysis/metabolism
MH  - Lymphocyte Activation/immunology
MH  - Neoplasms/*diagnosis/immunology
MH  - Orthomyxoviridae/immunology
MH  - Perforin/*analysis/metabolism
MH  - Staining and Labeling
MH  - T-Lymphocytes, Cytotoxic/cytology/*immunology
EDAT- 2014/08/26 06:00
MHDA- 2015/05/12 06:00
CRDT- 2014/08/24 06:00
PHST- 2014/08/24 06:00 [entrez]
PHST- 2014/08/26 06:00 [pubmed]
PHST- 2015/05/12 06:00 [medline]
AID - 10.1007/978-1-4939-1158-5_6 [doi]
PST - ppublish
SO  - Methods Mol Biol. 2014;1186:75-86. doi: 10.1007/978-1-4939-1158-5_6.