GSM5057746: 2A: vax010_062_d21_L4; Homo sapiens; ATAC-seq
Cryopreserved PBMCs were thawed and innate immune cell subsets were isolated using FACS. Within the live gated cells, CD14+ monocytes were identified as CD14+ (fraction A) while a mixture of the remaining monocyte and dendritic cell subsets was identified as CD14-CD56-HLA-DR+ (fraction B). After sorting and depending on the number of isolated cells, fraction A and B were mixed at a 2:1 ratio to yield a solution of monocytes and dendritic cells enriched for CD14- cells. FACS-purified cells were processed for single nuclei ATAC-seq according to the manufacturer's instructions (10x Genomics, CG000168 Rev D). Briefly, nuclei were obtained by incubating PBMCs for 3.20 minutes in freshly prepared Lysis buffer following manufacturer's instructions for Low Cell Input Nuclei Isolation (10x Genomics, CG000169 Rev C). Nuclei were washed and resuspended in chilled diluted nuclei buffer (10x Genomics, 2000153). Next, nuclei were subjected to transposition for 1h at 37C on the C1000 touch PCR instrument (BioRad) prior to single nucleus capture on the 10x Chromium instrument. Samples were subjected to post GEM cleanup, sample index PCR, cleanup and library QC prior to sequencing according to the protocol. Samples were pooled, quantified and sequenced on NovaSeq 6000 instrument (Illumina) with at least minimum recommended read depth (25000 read pairs/nucleus).