1.4 numerical aperture objective 90

For performing IFAs, smears were prepared on Teflon-coated slides and fixed with 4% paraformaldehyde-0.0025% glutaraldehyde solution for 30 min. Slides were kept in a humidity chamber for each step. Fixed parasites were washed twice with phosphate-buffered saline (PBS) followed by permeabilization with 0.1% Triton X-100-PBS solution for 10 min. Parasites were washed with PBS and blocked with 3% bovine serum albumin (BSA)-PBS for 45 min at RT. Primary antisera prepared in 3% BSA-PBS was added to the parasites, and slides were incubated at 4°C. Antigens were visualized using anti-species secondary antibodies conjugated to Alexa Fluor. Images were obtained using a 100× 1.4-numerical-aperture (NA) objective 90° (Olympus) on a Delta Vision Elite high-resolution microscope (GE Healthcare Life Sciences).

For performing IFAs, smears were prepared on Teflon-coated slides and fixed with 4% paraformaldehyde-0.0025% glutaraldehyde solution for 30 min. Slides were kept in a humidity chamber for each step. Fixed parasites were washed twice with phosphate-buffered saline (PBS) followed by permeabilization with 0.1% Triton X-100-PBS solution for 10 min. Parasites were washed with PBS and blocked with 3% bovine serum albumin (BSA)-PBS for 45 min at RT. Primary antisera prepared in 3% BSA-PBS was added to the parasites, and slides were incubated at 4°C. Antigens were visualized using anti-species secondary antibodies conjugated to Alexa Fluor. Images were obtained using a 100× 1.4-numerical-aperture (NA) objective 90° (Olympus) on a Delta Vision Elite high-resolution microscope (GE Healthcare Life Sciences).