Figure 1
Experimental procedure and classificaiton of cells.
(a) Donor mice (n = 2) were infected with transgenic PbA-GFPpos parasites 8 days prior to the beginning of the experiment. On day 0 of experiment, blood was taken from the donor mice by cardiac puncture, pooled together and fluroescently-labelled with the red blood cell (RBC) stain DDAO-SE. These labelled RBCs (donor RBCs) were then transferred into the two groups of mice, a group of uninfected (naïve) mice and a group of mice that had been infected 5 days prior to infection with GFPneg PbA parasites. (b) Flow cytometry allowed RBCs to be distinguished from other cells based on size and structural complexity, using the measures of forward scatter (FSC) and side scatter (SSC). Doublets (two cells either adhering together or detected simulatenously) were excluded via FSC-Height (FSC-H) versus FSC-Area (FSC-A) plots. RBCs and parasites from the donor mice were distinguished from RBCs and parasites from the recipient mice by gating on GFP expression and DDAO-SE staining. Further, blood samples from mice were stained with Hoechst and Syto 84. This staining allowed late-stages parasites to be distinguished from early-stage parasites.
