Quantifying Parasite Sequestration In Mice

Mice were infected with a luciferase-expressing strain of P. chabaudi (PccASluc [45 (link)]) as described above but kept on a reverse light cycle, as sequestration occurs during the dark cycle [45 (link)]. At the time of maximum sequestration (12.00 – 14.00 h coordinated universal time (UCT), reverse light) mice were sacrificed and perfused systemically by injection of 10 ml PBS into the heart. Organs were harvested and 0.1 g of tissue was transferred to a Precellys homogenizer tube in PBS and dissociated for one cycle, 10 seconds at 4500 rpm in a Precellys Evolution Homogenizer. The sample was then diluted 1:10 in PBS, and an equal volume (100 μL) of Bright-Glo substrate (Promega) was added. Luciferase activity was measured after 2 min incubation using a Perkin Elmer VICTOR X Light Multilabel Plate Reader.