biostudies-literatureHu HNorwegian Research CouncilSwiss National Science FoundationEuropean UnionTrond Mohn stiftelseMinistry of Health and Care ServicesSNSF Bridgepgaf006https://www.ebi.ac.uk/biostudies/studies/S-EPMC11759286biostudies-literatureUnknown4(1)Here, we present Link-Seq, a highly efficient droplet microfluidic method for combined sequencing of antibody-encoding genes and the transcriptome of individual B cells at large scale. The method is based on 3' barcoding of the transcriptome and subsequent single-molecule PCR in droplets, which freely shift the barcode along specific gene regions, such as the antibody heavy- and light-chain genes. Using the immune repertoire of COVID-19 patients and healthy donors as a model system, we obtain up to 91.7% correctly paired immunoglobulin heavy and light chains. Furthermore, we map the V(D)J usage and obtain sensitivities comparable with the current gold-standard 10× Genomics commercial systems while offering full flexibility in experimental setup and significant cost savings. A further uniquPNAS nexusTargeted barcoding of variable antibody domains and individual transcriptomes of the human B-cell repertoire using Link-Seq.PMC11759286H2020 101037867 VaccelerateH2020 874866 INCENTIVEF-1162821872140B2-0_218721TMS2020TMT05284930EU IMI 2 101007799 Inno4VacBrokstad KAHu HKolmar LMa XZhou FGirardot CCox RJBenes VMerten CAfalseTargeted barcoding of variable antibody domains and individual transcriptomes of the human B-cell repertoire using Link-Seq.Here, we present Link-Seq, a highly efficient droplet microfluidic method for combined sequencing of antibody-encoding genes and the transcriptome of individual B cells at large scale. The method is based on 3' barcoding of the transcriptome and subsequent single-molecule PCR in droplets, which freely shift the barcode along specific gene regions, such as the antibody heavy- and light-chain genes. Using the immune repertoire of COVID-19 patients and healthy donors as a model system, we obtain up to 91.7% correctly paired immunoglobulin heavy and light chains. Furthermore, we map the V(D)J usage and obtain sensitivities comparable with the current gold-standard 10× Genomics commercial systems while offering full flexibility in experimental setup and significant cost savings. A further uniqu2025-01-01T00:00:00Z2025 Jan2025-04-05T10:18:13.159Z2025-04-05T10:18:13.159ZS-EPMC117592863986766810.1093/pnasnexus/pgaf006