Tissues must be processed into a monodispersion of cells, typically by mechanical maceration or douncing. Enzymatic digestion (e.g. collagenase, dispase) can be used for connective tissue and Trypsin may be used for macro-dissected tissues (monitor dissolution to single cells). Avoid extensive enzymatic digestion and processing agents containing glucosaminidase, amidase, neuraminidase, or mannosidase to prevent interference with ConA bead binding. Once a monodispersion is achieved, proceed to initial cell count.
Harvesting viable, monodispersed cells from tissues can be challenging; in some cases (especially when performing CUT&Tag) nuclei may be preferable. See literature, including the following papers, for additional methods:
Carpenter et al. Cell-type specific profiling of histone post-translational modifications in the adult mouse striatum. Nature Communications 13 (2022).
de Bock et al. HOXA9 cooperates with activated JAK/STAT signaling to drive leukemia development. Cancer Discov 8, 616-631 (2018).
Janssens et al. Automated in situ chromatin profiling efficiently resolves cell types and gene regulatory programs. Epigenetics Chromatin 11 (2018).
Larsen et al. Establishment, maintenance, and recall of inflammatory memory. Cell Stem Cell 28, 1758-1774 (2021).
Liu et al. Direct promoter repression by BCL11A controls the fetal to adult hemoglobin switch. Cell 173, 430-442 (2018).
Miao et al. Glucose dissociates DDX21 dimers to regulate mRNA splicing and tissue differentiation. Cell 186, 80-97 (2023).
Uyehara & McKay. Direct and widespread role for the nuclear receptor EcR in mediating the response to ecdysone in Drosophila. Proc Natl Acad Sci USA 116, 9893-9902 (2019).