Dumont 5 45

Mice were euthanized by ketamine hydrochloride and xylazine hydrochloride. Blood was collected by cardiac puncture. Perfusion was performed from the left ventricle with 10 ml 5 mM EDTA buffer, 20 ml PBS and 20 ml FACS buffer. RLNs were collected under a dissecting microscope. To collect plaques and ATLOs, we followed the method described previously^{7 (link),20 (link)}. In short, adipose tissue and paraaortic lymph nodes were carefully removed. The whole aorta was dissected and collected in cell culture dishes with pre-cooled FACS buffer. The aorta was opened in the longitudinal direction, and the plaque tissues were carefully removed using curved forceps (Dumont 5/45, Fine Science Tools) under the dissecting microscope. The remaining aorta was collected as ATLOs. Three separated cohorts of mice were used: (1) pools of three Apoe^−/− and three WT mice were used to collect plaques, ATLOs and RLNs, and pools of five Apoe^−/− and five WT mice were used to collect blood in the first cohort; (2) cell hashtags (hashtags, BioLegend, TotalSeq C) were used to label four Apoe^−/− and four WT mice individually to collect ATLOs and RLNs; and (3) pools of five Apoe^−/− mice were used to collect plaque tissues.