Human TCR profiling and differential expression on the ICELL8 cx system
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 cx system. As the name suggests, the workflow allows generation of libraries that provide information on both alpha- and beta-chain diversity.
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 cx 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 cx 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 ICELL8 cx 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.
Overview
- Sensitivity—RACE-based approach allows for the detection of low-abundance TCR variants
- Specificity—Full length reads, with majority of reads on target and accurate pairing information
- Differential expression by 5' sequence enrichment—generate differential expression data for all transcripts by mapping the 5' ends
- Flexible, high-throughput single-cell analysis—generate sequencing data from hundreds of single cells in parallel and analyze cells from up to eight different samples in a single experiment
More Information
Applications
- Human TCR repertoire analysis in single cells (TCRα and TCRβ clonotypes)
- Differential expression by 5' end capture
Components
The following items are required to complete the workflow:
| ICELL8 cx Single-Cell System (Cat. # 640188 & Cat. # 640189) | ||
|---|---|---|
| Cat. # | Product | 1 item required per: |
| 640182 | ICELL8 Human TCR a/b Profiling Reagent Kit | 1 chip |
| 640179 | ICELL8 Human TCR a/b Profiling - Indexing Primer Set* | 1 chip |
| 640180 | ICELL8 Human TCR a/b Profiling - Indexing Primer Set* | 5 chips |
| 640181 | ICELL8 Human TCR a/b Profiling - Indexing Primer Set* | 10 chips |
| 640200 | ICELL8 cx TCR Chip | 1 run |
| 640212 | ICELL8 Collection Kit – L | 1 chip |
| 640197 | ICELL8 cx Loading Kit | 1 chip |
| 640192 | ICELL8 384-Well Source Plate and Seal (5/pack) | 1 chip |
| 740609.50 | NucleoSpin Gel and PCR Clean-Up | 1 chip |
| 635011 | SMARTer-Seq Magnetic Separator - PCR Strip (or equivalent) | Reusable |
*Note: Cat. No. 640179 contains enough primers for one chip. If you plan on processing multiple chips at a time, please purchase the appropriate indexing primer set. Cat. No. 640180 has indexing primers for up to five chips, while Cat. No. 640181 has primers for up to 10 chips.
Additional product information
Please see the product's Certificate of Analysis for information about storage conditions, product components, and technical specifications. Please see the Kit Components List to determine kit components. Certificates of Analysis and Kit Components Lists are located under the Documents tab.
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