genomic DNA samples, we compared profiles from the same sorted T or B cells (1K-50K) from tumor metastatic biopsy samples generated using our DriverMap AIR assay, which employs mRNA as the template, with Adaptive Technology’s genomic DNA-based assay. To assess the difference between Immune Repertoire profiling of RNA vs. Moreover, repertoire analysis using genomic DNA as a template doesn’t identify the Ig isotype since the V(D)J and C regions on this isotype are separated from each other by an intron that prevents them from being amplified together by PCR. Using high-throughput phage-display immunoprecipitation sequencing (PhIP-Seq), we identified antibodies against 344,000 antimicrobial, immune, and food antigens in 497 individuals with IBD compared with 1,326 controls.
As a result, RNA-based immune receptor repertoire is usually dominated by a low number of abundant clonotypes and many of these abundant clonotypes could be antigen-induced or disease-specific.Īnother benefit of using mRNA over genomic DNA starting material is that using RNA as a template limits amplification to only “functional” expressed receptor chains rather than non-functional genes–such as pseudogenes and ORFs–which reduces background in the NGS data. A comprehensive overview of an IBD-specific antibody epitope repertoire is, however, lacking. Nat Commun 9:561 10.Blondel VD, Guillaume J-L, Lambiotte R. Also, the clonotypes express receptor mRNA at different levels, and activation of adaptive immunity induces significant up-regulation of TCR and BCR transcription in antigen-specific clonotypes (e.g., up to 1,000-fold for plasma B cells). Marcou Q, Mora T, Walczak AM (2018) Highthroughput immune repertoire analysis with IGoR.
The higher number of starting RNA template copies has been shown to significantly increase the detection level of T-cell receptors. Additionally, pairing the clonal information with cellular phenotype can identify disease-relevant cell types that can be insightful for disease prognosis and to develop targeted immunotherapy strategies.Īlthough both genomic DNA and RNA can be used for AIR Profiling, the copy number of mRNA per cell is at least 10- to 100-fold more than gDNA. For example, tracking TCR/BCR diversity in patients across time points can reveal clonal dynamics that correlate with treatment response or other clinically relevant features. This results in a diverse repertoire called the adaptive immune repertoire (AIR) that comprises multiple individual clonotypes (sequence) for particular receptor chains.Īdaptive Immune Repertoire (AIR) Profiling is essential for designing effective therapeutic strategies for immune-mediated diseases. Diversity among B cell and T cell receptors is primarily produced by V(D)J recombination, which involves the shuffling and joining of the variable (V), diversity (D), joining (J), and constant region (C) gene segments. Adaptive immunity relies on B and T cells that recognize foreign antigens via hypervariable B cell and T cell receptors (BCRs and TCRs).
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