Differences between ELISPOT assays and other approaches for measuring antigen-specific T cell immunity
Every naïve T cell expresses a unique T cell receptor (TCR) which is specific for a single antigen. In order to be able to recognize potential infinite numbers of microbial antigens and at the same time to distinguish them from self antigens, T cells system relies on an close to infinite number (~1012) of different possible T cell specificities.
Subsequently, the frequencies of T cells recognizing individual antigens are very low. As a result of the clonal expansion associated with infections, the frequency of antigen-specific effector cells can transiently rise to as high as 1:100, but these frequencies typically settle in the range of 1:10,000 for effector memory or memory cells. The frequencies of specific T cells in blood after immunizations with subunit vaccines (proteins/peptides), environmental sensitization and in long-term, anti-microbial immunity rarely exceed the 1:10,000 thresholds.
Therefore, techniques that measure T cell immunity need to be sensitive enough to reliably detect such rare cells. In addition to measuring the frequencies of antigen specific T cells, which delineates the magnitude of immunity, it is essential to understand the functions of these T cells. The quality of T cell immunity depends on whether these cells are Th1 or Th2 (Tc1/Tc2) type cells and whether they are capable of direct killing. Sophisticated assays therefore will also provide information on the effector lineage of the antigen-specific T cells.
Optimized ELISPOT assays measure the magnitude and the quality of T cell immunity at single-cell resolution by detecting individual events of antigen-specific T cells that engage in secretion of cytokines and effector molecules such as granzyme B and/or perforin. The following is a comparison of the performance and utility of ELISPOT assays with other T cell diagnostic techniques including bulk supernatant measurements, Tetramer/Pentamer, and ICS techniques.
Interested in learning more about the advantages and applications of ELISPOT? Download Applications and Advantages of ELISPOT