Guidance on Processing the 10x Genomics Single Cell Gene Expression Assay.

The demand for technologies that allow the study of gene expression at single cell resolution continues to increase. One such assay was launched in 2016 by the US-based company 10x Genomics Inc. Utilizing the power of the single cell on a large scale (Zheng et al. Nat Commun 8:14049, 2017)-capturing thousands of cells at once-has shaped life sciences ever since and allowed researchers to discover new insights within their respective fields of study such as oncology, neurobiology, and immunology (among others). Obtaining high-data quality is the key to being able to make these meaningful discoveries, which in turn is directly linked to the quality of the initial cell (or nuclei) suspension that is used to load the 10x Genomics Chromium Single Cell Gene Expression assay. A successful workflow relies on a cell suspension which is fully dissociated, extremely clean, and of high viability. While the workflow itself has been detailed elsewhere (De Simone et al. Methods Mol Biol 1979:87-110, 2019), in this chapter we will focus on the importance of the quality of the initial cell suspension, as well as common mistakes that can occur while running a Single Cell Gene Expression assay. The descriptions of these tips and tricks refer to the current version of the 10x Genomics User Guide (Chromium Single Cell 3' Reagent Kits User Guide (v3.1 Chemistry Dual Index). https://support10xgenomicscom/single-cell-gene-expression/index/doc/user-guide-chromium-single-cell-3-reagent-kits-user-guide-v31-chemistry-dual-index) which can be downloaded from the Support section on the 10x Genomics website (10x Genomics website. https://www10xgenomicscom). These documents and user guides are continuously improved and updated; hence, it is important to regularly check the company's website for the most recent version.