Cb 300 k2e

Parasitemia was measured by blood smear or flow cytometry with similar results. For flow cytometry, 3 μl of tail blood was collected using an EDTA-coated Microvette (CB 300 K2E; Sarstedt) and diluted in 500 ml PBS. The diluted blood was labeled with antibodies directed against Ter119 (fluorescein isothiocyanate [FITC], TER-119, 1/30; Miltenyi Biotec), CD71 (phycoerythrin [PE], C2, 1/300; BD Pharmingen), and CD41‐PE‐Cy7 (MWReg30, 1/100; BioLegend), fixed, and permeabilized with 4% paraformaldehyde (PFA) with 0.6% saponin for 10 min, followed by 4′,6-diamidino-2-phenylindole (DAPI) staining.

After an overnight fast, 300μL of capillary blood (obtained from finger pricks) was collected into Microvette® capillary blood collection tubes treated with di Potassium EDTA (CB 300 K2E, Sarstedt, Germany) and kept on ice immediately. The blood samples were transferred and stored at −80°C until DNA extraction [23 (link)].

Male 14 to 18-week-old Ppara mice were placed in the calorimetric cages (Labmaster, TSE, BadHomburg, Germany) to measure food intake, locomotor activity, oxygen consumption and CO₂ production during seven days; energy expenditure was normalized to lean mass. For acute fat load test, mice were fasted for 3 h and gavaged with 100 μl of olive oil (Sigma-Aldrich). Blood samples during oral and intraperitoneal glucose tolerance tests (OGTT, IPGTT) and acute fat load test were collected from the tail vein in EDTA coated tubes (Sarstedt, CB 300 K2E). For OGTT or IPGTT, mice were fasted 12 h overnight and gavaged or injected, respectively, with 2 mg of glucose per g of body weight. For bomb calorimetry of the faeces, 48-hour faeces were collected from the cage bedding (2 mice per cage), vacuum dried, pulverized to the fine powder and analysed in the calorimeter (Parr, 6100, USA). Consumed energy was calculated by formula: digestible energy (kcal) = food intake (g) x food caloric density (kcal/g) − faeces weight (g) x faeces caloric density (kcal/g).