Fig. 1

Overview of the Cell Hashing workflow. A Sample collection and distribution overview. Cell Hashing allows flexible sample collection, including collection of multiple samples over time from the same donor. Each sample can then be thawed and stained with antibodies conjugated to Hash Tag Oligos (HTOs), each of which contains a unique barcode sequence. Once stained, samples can be mixed and distributed across wells for processing to reduce batch effects. B BarWare pipeline overview. After sequencing, two libraries are generated for each 10x Genomics Chromium well: a RNA library, and a HTO library. RNA libraries are aligned and converted UMI count matrices containing cell x gene counts (top), and HTO libraries are counted by Barcounter to generate cell x HTO counts (bottom). BarMixer is then used to convert HTO counts to sample assignments for each cell. BarMixer then combines sample assignments with the scRNA-seq to split sample data within each well, and finally merges data from each sample from all wells to generate sample-specific output. CellBC, Cell Barcode; UMI, Unique Molecular Identifier; HTO, Hash Tag Oligonucleotide