description: A bead-based strategy to screen viable CD45- cells was applied. The whole mouse aortae (n=3) of the Western diet group were isolated after intracardiac perfusion with ice-cold PBS. The finely cut aortae were incubated in 1x aortic dissociation enzyme cocktail (450 U/mL collagenase type I, 60 U/mL DNase I, and 60U/mL hyaluronidase) at 37oC for 1 h. The digestion was halted by adding FACS buffer (2 mM EDTA, 2% FBS). The cell suspension was transferred through 50 m cell strainer for subsequent incubation with dead cell removal microbeads (Miltenyi Biotec, North Rhine-Westphalia, Germany) at room temperature for 15 min. Viable cells were collected by transferring cell suspension through a MS column (Miltenyi Biotec) on a magnet. Later, the viable cells were incubated with CD45 microbeads (Miltenyi Biotec) on ice for 15 min for negative selection through a LS column (Miltenyi Biotec) on a magnet. The flow-through cells were centrifuged and resuspended in 0.04% BSA-PBS for droplet-based scRNA-seq.
attributes.tag: strain
attributes.value: ApoE-/-
attributes.tag: age
attributes.value: 10 weeks
attributes.tag: sex
attributes.value: male
attributes.tag: tissue
attributes.value: Aortae
attributes.tag: sample_type
attributes.value: Single cell
attributes.tag: treatment
attributes.value: Western diet
attributes.tag: BioSampleModel
attributes.value: Model organism or animal