SCITO-seq2: ultra-high-throughput single-cell transcriptome and epitope sequencing.

We introduce SCITO-seq2, an enhanced successor to SCITO-seq that integrates probe-based RNA detection with the established ultra-high-throughput protein profiling. SCITO-seq2 achieves robust quantification of transcripts and surface proteins across more than 100,000 cells, with a shared pool barcoding strategy ensuring precise matching of molecular profiles within multiplexed droplets. SCITO-seq2 is compatible with cell hashing technology, allowing efficient sample multiplexing. We demonstrate its utility in autoimmune diseases, including childhood systemic lupus erythematosus and CTLA4 haploinsufficiency with autoimmune infiltration, enabling the detection of minor immune clusters and disease-specific protein signatures. This platform establishes a scalable, streamlined, and cost-effective next-generation single-cell multi-omics workflow.