This protocol describes nuclei extraction from snap frozen tissue (human or mouse) using Singulator, including washing nuclei suspension in buffer supplemented with sucrose, and nuclei FACS sorting. The resulting nuclei suspension is suitable for scRNA-seq using a platform of choice (as 10X Chromium, or inDrops). The protocol has been validated on various snap frozen human and mouse tissues (lung, brain, prostate, tumors, core needle biopsies amongst others).