4. Single-Cell Immune Profiling Service

In addition to bulk AIR profiling (kit and services), Cellecta offers single-cell T and B immune profiling custom service using the 10x Genomics Chromium V(D)J assay. The Chromium Single Cell 5’ Library Construction Kit and V(D)J Enrichment Kit offer comprehensive, scalable solutions for measuring gene expression and immune repertoire information from the same cell. This protocol can generate an enriched T-cell library, an enriched B-cell library, and a 5’ Gene Expression library from amplified cDNA from the same cells.

This service can profile full-length (5’ UTR to constant region), paired T-cell receptor (TCR), or B-cell immunoglobulin (Ig) transcripts from 100-10,000 individual cells per sample. A pool of ~750,000 barcodes is sampled separately to index each cell’s transcriptome. It does so by partitioning thousands of cells into nanoliter-scale Gel Beads-in-emulsion (GEMs), where all generated cDNAs share a common 10x Barcode. Libraries are generated and sequenced, and 10x Barcodes associate individual reads back to the individual partitions. Detailed information about experimental design, sample preparation, and downstream data analysis may be found at: www.10xgenomics.com/support/single-cell-immune-profiling.

Single-cell V(D)J immune profiling and data analysis services are available for cryopreserved (in DMSO) lymphocyte cell samples (e.g., PBMC or sorted immune cells). For single-cell AIR experiments, it is desirable to use samples in which lymphocytes are enriched by centrifugation (PBMC), magnetic beads, or FACS sorting. It is recommended to use low-pressure sorting (if possible, or a nozzle > 100 um). It is also highly recommended for all immune cell samples undergo viability testing using live-dead staining to confirm that at least 70% of cells are alive. Upon receiving the cells, they will be processed using the Chromium controller, and 5’-RNA expression profiling libraries will be constructed using the RNA V(D)J protocol. The samples will be sequenced and analyzed using Cell Ranger V(D)J and Loupe V(D)J Browser tools.