This is a summary, written by members of the CITF Secretariat, of:
Law JC, Watts TH. Considerations for choosing T Cell assays during a pandemic. J Immunol. 2023 Jul 15;211(2):169-174. https://doi.org/ doi: 10.4049/jimmunol.2300129.
The results and/or conclusions contained in the research do not necessarily reflect the views of all CITF members.
A CITF-funded study, published in The Journal of Immunology, reviewed the benefits and limitations of commonly used methods for assessing T cell immunity across SARS-CoV-2 studies. These include ELISPOTReports the frequency of cytokine-secreting cells by capturing cytokines released during stimulation with plate-bound antibodies, intracellular cytokine staining (ICS)Reports the functionality of antigen-specific T cells by determining the frequency of cytokine-producing cells following antigen stimulation, activation induced markers (AIM)Identifies T cell responses based on the upregulation of several different activation markers upon antigen restimulation, cytokine secretion assays (CSA)This simultaneously quantifies multiple cytokines, up to 50 depending on the company, secreted into cell culture supernatants after antigen stimulation, and peptide MHC tetramer staining (pMHC)The ability to directly probe epitope-specific CD4 and CD8 T cells without the need for prior stimulation, thus providing a more accurate estimate of ex vivo epitope specific T cell frequencies, including exhausted cells. The authors highlighted the necessity for ongoing efforts to standardize T cell assays to ensure consistency and comparability across research studies. This study was led by Dr. Tania Watts (University of Toronto).
- Various T cell assays address different aspects of T cell function and activation state, with the consequence that results reflect the heterogeneity and wide range of T cell responses.
- T cell assays pose challenges due to variations in in vitro culture conditions, sample processing, stimulation methods, and diverse markers used in flow cytometry panelsA technology that provides rapid multi-parametric analysis of single cells in solution.
- To detect antigen specific T cells, AIM and ICS assays are the most frequently used flow-based assays for clinical studies – they can detect multiple cell surface markers and cytokines.
- To analyze T cell function, ELISPOT, CSA, and ICS are more frequently used.
- The choice of T cell assay depends on the research question and the sample size at hand. Simple and scalable assays like ELISPOT, multiplexed CSA, or AIM assays are ideal for rapid deployment in larger study cohorts, while more in-depth assays like ICS or single cell transcriptomicsDetermining mRNAs are expressed in individual cells following antigen stimulation may be suitable for detailed characterization.
In conclusion, the review underscores the importance of selecting the most appropriate assay for accurately measuring T cell responses, balancing the ease of use versus the depth of information required. Standardization of assays for detecting antigen-specific T cell responses is crucial for reliable and comparable results across different laboratories that are evaluating the immunogenicity of novel therapeutics and vaccines in a pandemic.