Fig. 1

Cell preparation for droplet-based single-cell transcriptional profiling. Schematic of experimental workflow. Cultured human (HEK) and mouse (3T3) cells were dissociated, mixed and further processed to analyse the transcriptomes of either live or fixed cells by Drop-seq. Washed cells were gently resuspended in 2 volumes of ice-cold PBS, then fixed by adding 8 volumes of ice-cold methanol. Methanol-fixed cells could be stored for up to several weeks at –80 °C. Prior to Drop-seq, cells were washed before passing them through a 35- to 40-μm cell strainer. Cells were then separately encapsulated in droplets together with a single bead in a microfluidic co-flow device and single-cell transcriptomes sequenced in a highly parallel manner. Downstream analysis and systematic quantitative comparisons were subsequently made from separate experiments using live or fixed cellular input material with an R package ('dropbead') that we developed and is freely available for download