Visualizing Cryptococcus neoformans Capsule in Brain Tissue

C. neoformans cells and CPS released in tissue were stained using GXM-specific mAb 18B7. Slides were blocked, and mAb 18B7 (2 μg/ml) was added for 1 h at 37°C. After the slides were washed, fluorescein isothiocyanate (FITC)-conjugated GAM Ab [dilution, 1:250; 1% BSA] was applied for 1 h at RT. Neurons in tissue sections were stained with DAPI (4′,6-diamidino-2-phenylindole) and microtubule-associated protein-2 (MAP-2) as described previously [74 (link)]. Microscopic examinations of brain sections were performed with a fully motorized Carl Zeiss Axio Observer Z1 confocal microscope. Confocal images of blue, green, and red fluorescence were conceived simultaneously using a multichannel mode. Z-stack images and measurements were corrected utilizing Zen software in deconvolution mode.