Our high-throughput TCR profiling solution leverages SMART technology (Switching Mechanism at 5" End of RNA Template) and employs a 5" RACE-like approach to capture complete V(D)J variable regions of TCR transcripts from hundreds of single cells isolated on the ICELL8 Single-Cell System. As the name suggests, the workflow allows generation of libraries that provide information on both alpha- and beta-chain diversity.

The workflow incorporates Illumina-specific adapter sequences during cDNA amplification for the generation of indexed libraries that are ready for sequencing on Illumina platforms. The application also enables you to perform differential expression analysis by 5" end capture in parallel.

Single T cells isolated with the ICELL8 system, as well as positive and negative control samples, are dispensed into a 5,184-nanowell TCR chip that contains preprinted barcodes. Each TCR chip contains 1,728 unique barcodes, and each barcode is printed three times on the chip. Following identification of single-cell-bearing wells using the ICELL8 CellSelect Software, SMART RT-PCR reagents are dispensed to the wells of samples chosen for further processing. Barcoded cDNAs from the selected wells are PCR-amplified in-chip. The full-length barcoded amplicons are then pooled off-chip, and the purified cDNA is used as template in the first TCR-specific PCR. The product from the first TCR-specific PCR is used as a template in a second TCR-specific PCR to incorporate Illumina indexed primers.