Hc pl apo 63 1.40 oil objective

For P. berghei liver stage parasites, 100,000 HeLa cells were seeded in glass-bottomed imaging dishes. Salivary glands of female A. stephensi mosquitoes infected with CEN4-GFP parasites were isolated and disrupted using a pestle to release sporozoites, which were pipetted gently onto the seeded HeLa cells and incubated at 37°C in 5% CO₂ in complete minimum Eagle's medium containing 2.5 μg/ml amphotericin B (PAA). Medium was changed 3 h after initial infection and once a day thereafter. For live cell imaging, Hoechst 33342 (Molecular Probes) was added to a final concentration of 1 μg/ml, and parasites were imaged at 24, 48, 55 h post-infection using a Leica TCS SP8 confocal microscope with the HC PL APO 63×/1.40 oil objective and the Leica Application Suite X software.

To study localisation of NDC80–GFP in P. berghei liver stages, 100,000 HeLa cells were seeded in glass-bottomed imaging dishes. Salivary glands of female A. stephensi mosquitoes infected with NDC80–GFP parasites were isolated and sporozoites were released using a pestle to disrupt salivary gland cells. The released sporozoites were pipetted gently onto the HeLa cells and incubated at 37°C in 5% CO₂ in air, in complete minimum Eagle's medium (Thermo Fisher Scientific) containing 2.5 μg/ml amphotericin B. For initial infection, medium was changed at 3 h post-infection and thereafter once a day. To perform live-cell imaging, Hoechst 33342 (Molecular Probes) was added (1 μg/ml) and imaging was done at 55 h post-infection using a Leica TCS SP8 confocal microscope with the HC PL APO 63×/1.40 oil objective and the Leica Application Suite X software.

HeLa cells (1 × 10⁵ cells) were seeded in glass‐bottomed imaging dishes. Salivary glands of female A. stephensi mosquitoes infected with myosin‐GFP parasites were isolated and disrupted using a pestle to release sporozoites, which were pipetted gently onto the seeded HeLa cells and incubated at 37°C in complete minimum Eagle's medium containing 2.5 μg/ml amphotericin B (PAA) and gassed with 5% CO₂ in air. Medium was changed 3 hr after initial infection and once a day thereafter. For live cell imaging, Hoechst 33342 (Molecular Probes) was added to a final concentration of 1 μg/ml, and parasites were imaged at 24, 48, and 55 hr post infection using a Leica TCS SP8 confocal microscope with the HC PL APO 63×/1.40 oil objective and the Leica Application Suite X software.