Flow cytometric quantitation of oxidative product formation by heterophils from orange-winged Amazon parrots (Amazona amazonica amazonica)

Oxidative metabolic burst activity by heterophils from orange-winged Amazon parrots (Amazona amazonica amazonica) was indirectly evaluated by measuring the oxidation of non-fluorescent intracellular 2′,7′-dichlorofluorescin (DCFH) to fluorescent 2′,7′-dichlorofluorescein (DCF) with flow cytometry. Heterophils were isolated from whole blood samples by preconcentrating the white blood cells with 3% hetastarch before running the samples through a polysucrose and disodium diatrizoate centrifugation. Phorbol myristate acetate (PMA) was used to activate the heterophils. Results show that heterophils from orange-winged Amazon parrots can oxidise DCFH to a fluorescent product and a wide range of DCF production was observed among individual birds. The production of DCF increased with PMA concentrations and significant differences were seen between the highest and lowest concentrations of PMA. These results support the concept that heterophils from orange-winged Amazon parrots can produce oxidative metabolites and that these heterophils appear to have functional heterogenicity.