Fluorescence-activated nuclei sorting (FANS) on human post-mortem cortex tissue enabling the isolation of distinct neural cell populations for multiple omic profiling v2

Increased understanding of the functional complexity of the genome has led to growing recognition about the role of epigenetic/transcriptional variation in health and disease. Current analyses of the human brain, however, are limited by the use of “bulk” tissue, comprising a heterogeneous mix of different neural cell types. As epigenetic processes play a critical role in determining cell type-specific patterns of gene regulation it is important to consider cellular composition in regulatory genomic studies of human post-mortem tissue, and there is a need for methods to purify populations of specific cell-types. Furthermore, the valuable nature of human post-mortem tissue means it is important to use methods that maximize the amount of genomic data generated on each sample. This protocol describes a method that uses fluorescence-activated nuclei sorting (FANS) to isolate and profile nuclei from multiple different human brain cell-types from frozen post-mortem tissue. This protocol can be used to robustly purify populations of neuronal (NeuN+ve), oligodendrocytes (SOX10+ve), microglia (IRF8+ve) and other glial origin nuclei (NeuN-ve/SOX10-ve/IRF8-ve) from adult post-mortem frozen brain, with each tissue sample yielding purified populations of nuclei amenable to simultaneous analysis of i) DNA modifications (via bisulfite sequencing / array), ii) histone modifications (CUT&Tag), iii) open chromatin analysis (ATAC-seq), and iv) gene expression (snRNA-seq).