Encyclopedia of Experiments: Immunology
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Transcript
Begin with a fungal suspension of Cryptococcus. Add a solution of fungal-derived bioparticles conjugated with a pH-sensitive fluorescent dye to stain the Cryptococcus cells.
Wash to remove the unbound dye. Transfer the stained fungal suspension into a multi-well plate with adhered macrophages — or phagocytic immune cells.
Introduce the minimum inhibitory concentration of a drug, such as aspirin or ibuprofen, and incubate to facilitate the drug uptake.
The drug molecules interact with cyclooxygenase-2 — a prostaglandin-synthesizing enzyme, inhibiting its activity.
This prevents the conversion of arachidonic acid to prostaglandin, preserving intracellular cAMP levels and enhancing the macrophage phagocytic ability, facilitating the internalization of fungi.
The fungi-containing phagosome fuses with the lysosome — an acidic organelle, forming a phagolysosome with an acidic pH and allowing the dye to fluoresce.
Drug-treated cells exhibit a higher fluorescence than untreated cells, suggesting the drug’s efficacy in enhancing the phagocytic ability of macrophages.
