GSM3294727: 067_Lgate_CD3PosCD14Pos; Homo sapiens; RNA-Seq
Cryopreserved PBMC were thawed, stained with live/dead stain, anti-human CD14 and CD3 antibodies, and single cell sorted using BD Aria 3 cell sorter direclty in cell lysis buffer. Monocyte gate (Mgate) and lymphocyte gate (Lgate) were defined from FSC and SSC parameters. Monocytes were defined as live Mgate CD14+CD3- cells, T cells were defined as live Lgate CD14-CD3+ cells, CD14+CD3+ Mgate were defined as live Mgate CD14+CD3+ cells and CD14+CD3+ Lgate were defined as live Lgate CD14+CD3+ cells. Total RNA was extracted using Smart-seq2 method as previously described (Picelli et al., 2013). Single cell RNAseq was performed as described in Picelli et al.Nat Protoc. 2014, PMID:24385147. 0.3 to 0.5 ng of pre-amplified cDNA was used to generate barcoded Illumina sequencing libraries (Nextera XT library preparation kit, Illumina) in an 8µL reaction volume.
GEO Accession
GSM3294727