GSM4310101: Macrophages SPTA aCD40 3; Mus musculus; RNA-Seq
The abdominal cavity of a living, deeply anesthetized mouse was opened, and the portal vein was catheterized for in situ liver perfusion and digestion with 50 ml of 0.06 U/ml collagenase B buffered solution (Roche, 11088815001). Once digested, livers were dissected out and the mouse was sacrificed. The digested livers were then mechanically disaggregated in a petri dish on ice and filtered through a 100 μm pore cell strainer. The cell suspensions were centrifuged twice at 60 x g for 2 min at 4°C, and the pellets were discarded. The supernatants were then centrifuged at 300 x g for 5 min at 4°C to obtain a pellet of nonparenchymal liver cells containing liver macrophages. To select for F4/80+ macrophages from liver cell suspensions, Dynabeads Sheep anti-Rat IgG (Invitrogen, 11035), in combination with purified rat anti-mouse F4/80 antibody (BD Pharmingen, 0.5 mg/ml, 565409) ), and a DynaMag™-2 Magnet (Thermo Fisher Scientific, 12321D) were used according to the manufacturer's protocol (Invitrogen) RNeasy Micro kit (Qiagen, Hombrechtikon, Switzerland) according to the manufacturer's protocol, including on-column DNase I treatment. RNA samples using the Illumina TruSeq RNA stranded kit following the manufacturer's instructions.