The set of DriverMap™ Single-cell Adaptive Immune Receptor (scAIR) Chain Profiling Assays enables identification of TCR/BCR paired full-length (V(D)J) immune receptor sequences of the single T or B cells sorted in a 96-well plate. This assay allows a cost-effective strategy for scAIR repertoire analysis of hundreds of sorted single cells, selected based on the expression of specific markers (e.g., activated, dysfunctional, memory, etc.), or after labeling with specific antigen or MHC-peptide complexes. The paired full-length receptor sequence information is necessary to clone and express functional TCRs, BCRs, and antibodies for follow-up studies of receptor-antigen interaction and the development of cell-based or antibody-based therapies.
The DriverMap scAIR Chain Pairing Assays are designed to amplify in a multiplex RT-PCR reaction full-length (CDR1-CDR2-CDR3) regions for human, mouse or human-mouse hybrids of TCR or BCR chains, from T or B cells sorted in 96-well plates (s) and are available in several formats:
- The scAIR TCR Chain Pairing Assay (Human/Mouse) allows analysis of alpha-beta (TRB-TRA) chains of TCRs in sorted human or mouse T cells.
- The scAIR BCR Chain Pairing Assay (Human/Mouse) allows analysis of all isotypes of heavy-light (IGH, IGK, IGL) chains of BCRs in sorted human or mouse B cells.
- The scAIR TCR Chain Pairing Assay (Hybrid, Human-Mouse) allows analysis of human alpha-beta (TRB-TRA) V(D)J regions fused with mouse C region of hybrid TCRs in sorted mouse T cells.
- The scAIR BCR Chain Pairing Assay (Hybrid, Human-Mouse) allows analysis of human heavy-light (IGH, IGK, IGL) V(D)J regions fused with different isotypes of mouse C region of hybrid BCRs in sorted mouse B cells.
The amplified immune receptor cDNA products for all assays contain Validator Barcodes (VBCs), cell/well-specific barcodes, and P5 & P7 indexes compatible with Illumina’s Next-Generation Sequencing (NGS) to analyze paired immune receptor chain RNA sequences at single-cell resolution.
Key Features of the DriverMap scAIR Chain Pairing Assay:
• The reverse gene-specific primers (Rev GSPs) for immune receptors are designed with 10-nt cell/well-specific barcodes to pool 96 cells from each plate in a single test tube immediately after the initial Rev GSP-RNA hybridization step. This protocol design significantly simplifies the workflow, allows us to process several plates simultaneously, and reduces library preparation time for follow-up reverse transcription and PCR steps.
• The reverse GSPs are also designed with VBCs, which facilitate the accurate quantification of the copy number of cDNA molecules used in amplification steps and the selection of correct immune receptor chains from mutated sequences generated at amplification and NGS steps.
• The scAIR Chain Pairing assay works with FACS-sorted cells in a 96-well plate without needing special microfluidic equipment or complex split-pool barcoding workflows.
• The scAIR Chain Pairing employs several mechanisms to minimize off-target and primer dimer background product amplification.
• The kit contains control PBMC RNA that can be run in parallel for accurate quantitation. DriverMap Spike-in Controls can also be purchased to add to the single-cell experiment as additional controls.
• NGS data can be analyzed with open-source or MiXCR, Platforma, and RSEM software packages available online.
Related Products and Services:
- DriverMap™ AIR TCR/BCR Profiling Kit (Bulk) (human/mouse RNA/DNA)
- DriverMap™ EXP Human Genome-wide 19K Profiling Kit
- DriverMap™ Spike-in Controls
- DriverMap™ scAIR Immunophenotyping Assays (with immune markers)
Visit our website for scAIR Custom Services from Cellecta.
Please read the entire user manual before proceeding with your experiment.
PLEASE NOTE: The purchase of all Cellecta Products is covered by Cellecta’s standard Terms and Conditions of Sale as described on Cellecta’s website, and selected Products containing particular technology or having certain features are also subject to restrictions on use as outlined in the Label License section of our website. Please review these Terms and Label License Restrictions before opening and using your Product and, if you are not able to abide by the restriction, contact Cellecta to return the item to Cellecta for a full refund.
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