Skip to main content
. 2022 Oct 13;12:1004339. doi: 10.3389/fcimb.2022.1004339

Table 1.

Methods to examine the regulation of T cell repertoires by gut microbiota.

Methods Advantages Disadvantages
TCR-Tg cell transfer - Easier identification of Ag-specific T cells by using congenic marker
- Precise determination of Ag-specific T cell proliferation (CFSE or CTV labeled donor T cells)
- Examination of the interaction between Ag-specific T cells and other immune cells (i.e., B cells)
- Single TCR specificity
- Not reflect the physiological frequency of cognate Ag specific T cells
- Intra-clonal competition distorts natural Ag-specific T cell responses against microbiota.
- Limited use depending on HLA types
pMHC Tetramer staining - Direct examination and quantification of Ag-specific T cell frequency under steady state and inflammatory settings
- Examination of physiological Ag-specific T cell responses
- Relatively lower affinity between pMHC tetramer and TCR (limitation of visualization in flow cytometry)
- Limited use depending on HLA types
- Broad TCR specificities within pMHC tetramer+ T cells
Bulk TCR gene sequencing - Less expensive than single cell based TCR gene sequencing
- Relatively bigger sample size
- Lack of information on the TCR diversity created by TCR chain pairing*
- Limited association between TCR clonality and T cell subset/function (cytokine, transcription factor expression)**
Single cell-based TCR gene sequencing - Strong association with TCR clonality and T cell subset/function
- Possible examination of the TCR diversity created by TCR chain pairing
- Combination with single cell-based transcriptomics & other -omics
- Relatively expensive and limitation on sample size (typically less than 2-3 x 104)
- Requirement of complex analytical methods

*Assessment of TCR chain paring is not required in case of bulk TCR gene sequencing based on TCRα or TCRβ-restricted mice., **addressed by TCR sequencing on T cells sorted based on the transcription factor or cytokine reporters.