Ucth1

For sorting MuSK multimer–reactive B cells, B cells were enriched from cryopreserved PBMCs using negative selection beads (Stemcell Technologies). They were incubated with live/dead stain, then stained with 20 μg/ml MuSK multimer on ice for 30 minutes. Cells were then costained (using manufacturer’s recommended dilutions) with fluorescently labeled antibodies against CD3 (Invitrogen, Pacific orange; UCTH1), CD14 (Invitrogen, Pacific orange; TUK4), CD19 (BD Biosciences, PE Cy7; SJ25C1), CD27 (BD Biosciences, PE; M-T271), and CD38 (BD Biosciences, V450; HB7) before sorting on an FACSAria (BD Biosciences) instrument. For general B cell immunophenotyping, B cells were defined as live CD3^–CD14^–CD19⁺ cells, memory B cells as live CD3^–CD14^–CD19⁺CD27⁺CD38^– cells, and antibody-secreting cells (plasmablast phenotype) as CD3^–CD14^–CD19⁺CD27⁺CD38^hi.

B cells were enriched from cryopreserved PBMCs using negative selection beads (Stemcell Technologies; 19554). They were incubated with live/dead stain, then stained for 30 minutes on ice with fluorescently labeled antibodies against CD3 (BD Biosciences, V500; UCTH1), CD14 (Invitrogen, Pacific orange; TUK4), CD19 (BioLegend, PE Cy7; SJ25C1), CD27 (BD Biosciences, PE; M-T271), IgD (BD Biosciences, FITC; IA6-2), IgM (BioLegend, PerCP/Cyanine5.5; MHM-88) and CD38 (BioLegend, BV421; HB-7) using manufacturer’s recommended dilutions. The cells were sorted on an FACSAria (BD Biosciences) instrument and the population of CD3-CD14-CD19+IgD-CD27+IgM-CD38+ was collected for subsequent analysis by 10x Genomics. Sorted B cells were then loaded into the Chromium Controller (10 × Genomics). Single-cell gene expression libraries were prepared using the Chromium Single-cell 5′ Reagent Kit (10x Genomics; V 2.0) according to manufacturer’s instructions. Samples were sequenced on the NovaSeq 6000 Sequencing System (Illumina) with HiSeq paired-end, 150bp reads for 10 × Single cell BCR (BCR libraries) and 10 × Single cell 5 Prime (gene expression).