Dmi6000b af6000 microscope

The parasites were maintained in LIT medium as previously reported for fatty acid staining using BODIPY 500/510. After incubation, the cells were washed twice in PBS and placed on glass slides. The images were acquired with a digital DFC 365 FX camera coupled to a DMI6000B/AF6000 microscope (Leica). The images were analysed using ImageJ software.

Hemocytes were collected from non-infected and infected larvae and sampled on coverslips placed inside a 24 cell culture plate containing RPMI 1640, 10% FBS and 1% penicillin/streptomycin, and incubated at 30℃ in a CO₂ chamber. After 3 h, the samples were washed with IPS. The hemocyte aggregates were counted in a DMI6000B/AF6000 microscope coupled to a DFC365FX camera with the help of the LAS software for image capture (Leica).

CHO-K₁ cells (1.0 x 10⁴ per dish—Corning BioCoat Culture Dishes, New York, NY, United States), cultivated in RPMI medium supplemented with SFB 10% were incubated with CytoPainter LysoNIR Indicator Reagent (Abcam, Cambridge, United Kingdom—1:1000 –v/v) for 1 h, LysoNIR is a lysotropic dye that selectively accumulates in lysosomes via the lysosome pH gradient. After this time, the cultures were washed 2 times with PBS and infected with CDT forms (5 x 10⁵ per well– 50 parasites per cell) for three h. The cultures were washed another 2 times with PBS and incubated with MS (concentrations for IC₅₀ and IC₈₀ in the intracellular cycle– 20) and Hoechst 33342 (1:1000 –Thermo Fisher Scientific). After different incubation times (T0; 1 h, 6 h and 24 h), images were acquired with a digital DFC 365 FX camera coupled to a DMI6000B/AF6000 microscope (Leica), and Software AF6000 was used to estimate the percentage of parasites within parasitophorous vacuoles.