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. Author manuscript; available in PMC: 2014 Mar 28.
Published in final edited form as: Nat Protoc. 2011 Mar 31;6(4):553–561. doi: 10.1038/nprot.2011.313

Figure 3. Analysis of mCherry expression in transformed P.c. chabaudi.

Figure 3

(a) Immunofluorescent images of transformed mCherry-expressing P.c. chabaudi AS (mCherryPcc AS) and wild-type P.c. chabaudi AS (Pcc AS), demonstrating mCherry expression (red) and Hoechst labeling of DNA (blue) in the mosquito midgut (oocyst) and in each stage of the erythrocytic cycle in the mouse (ring, trophozoite, schizont). The top row shows an overlay of mCherry, Hoechst and bright-field images; the bottom row shows the mCherry image only. (b) Flow cytometry analysis of blood from mCherryPcc AS–infected mice, demonstrating mCherry expression (x axis) and Hoechst labeling of DNA (y axis). Three distinct populations of parasitized erythrocytes are gated based on their labeling with Hoechst, and the mean fluorescence intensity (MFI) of mCherry expression is shown for each population. This demonstrates that expression of the transgene increases with increasing DNA content and, therefore, parasite biomass (see Box 4 for details of flow cytometry analysis of blood). (c) Flow cytometry analysis of blood from uninfected (left), wild-type Pcc AS–infected (center) and transformed mCherryPcc AS–infected (right) mice, demonstrating mCherry expression (x axis) and Hoechst labeling of DNA (y axis). The MFI of mCherry expression in the Hoechst-positive population is shown above each graph. This demonstrates that all parasites (Hoechst-positive cells) are expressing the mCherry transgene (see Box 4 for details of flow cytometry analysis of blood). All animal experiments were carried out according to institutional guidelines (National Institute for Medical Research Ethical Review Panel) and UK Home Office regulations.