Fig. 3

Merged images of infected Vero cells with rickettsiae and comparison of four fixative, summary of results. We ran parallel experiments on Vero cells infected with two species of rickettsiae, namely R. akari and R. slovaca, for optimization of fixatives to properly preserve actin cytoskeleton. We tested four fixatives, 3.7% formaldehyde (FA), 4% paraformaldehyde (PFA), 4% paraformaldehyde in the cytoskeletal buffer (PFA + CB) and 4% paraformaldehyde in PHEM buffer (PFA-PHEM). Formaldehyde conserved actin as discontinuous filaments compared to PFA-based fixatives. Unspecific artifact observed in 4% PFA + CB treatment is marked with an arrow. Our results suggest that 4% PFA-PHEM buffer gives the most accurate image of the actin cytoskeleton and simultaneously allows monitoring rickettsiae using actin-based motility during infection inside the host cells. Scale bar represents 10 µm