Multiplexed single-cell gene expression and TCR sequencing data generated from human thymus and blood samples

Single-cell RNA sequencing (scRNA-seq) has improved our ability to study rare cell subsets. To maximise reagent utility, researchers often process samples from multiple donors or conditions simultaneously (multiplexing) and overload microfluidic chip channels (superloading). While superloading reduces processing time and costs, it increases the incidence of doublets, potentially complicating downstream analyses. To investigate the effects of superloading on primary immune cells, we generated and analysed single-cell gene expression and TCR data from standard- and super-loaded gel beads-in-emulsion (GEM) chip channels. While the transcriptomic profiles of the two loading methods were largely similar, we observed that most T cells expressing multiple TCR chains were doublets, underscoring the need for TCR configuration-based doublet removal for accurate T cell analysis.