2.6. NO production
The fluorescent probe 4-amino-5-methylamino-2′,7′-difluorofluorescein diacetate (DAF-FM DA, Molecular Probes) was used to
detect intracellular NO production. This probe is cell-permeant and
passively diffuses across cellular membranes. Once inside the cells,
it is deacetylated by intracellular esterases to become DAF-FM. After
reacting with NO, DAF-FM is further converted to DAF-FM triazole (λ-excitation=495,λemission=515) which exhibits about a 160-fold greaterfluorescence quantum efficiency. A volume of 200μL haemocyte
suspension was incubated with 10μM DAF-FM DA for 60 min at
room temperature in the dark. Then the fluorescence of cells was
recorded on FL1 detector of flow cytometer. Results were given as
mean of DAF-FMfluorescence, in arbitrary FL1 units.