MAITs from PBMCs and tonsils were single cell sorted using the Aria II cell sorter (BD Biosciences) directly into One Step RT-PCR reaction mix (NEB) loaded in MicroAmp Optical 96-well reaction plates (Applied Biosystems). MAITs were defined as CD3+ Vα7.2+ MR1-Tetramer+ cells and sorted based on CD161 expression. Following reverse transcription and preamplification reaction, a series of three nested PCR’s were run using primers for TCR sequence and gene expression as described (43 (link)). Subsequent sequencing data analysis was performed as described (43 (link)).
Protocol detail
Single-cell MAIT transcriptomic profiling
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Corresponding Organization : University of Utah
Other organizations : Primary Children's Hospital
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Variable analysis
independent variables
- Cell sorting: MAITs were defined as CD3+ Vα7.2+ MR1-Tetramer+ cells and sorted based on CD161 expression
- TCR sequence
- Gene expression
- Reverse transcription and preamplification reaction
- Three nested PCRs using primers for TCR sequence and gene expression
- Positive control: Not specified
- Negative control: Not specified
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