Example sentences for: hff
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Although the location of activated NADPH oxidase complex in phagolysosomes has been reported, the release of either H 2 O 2 or O 2 •- in situ upon stimulation with PMA has not been directly shown in live cells [ 10 11 ] . In order to directly measure the oxidative activity in lysosomes, I use a reduced fluorescein derivative that is conjugated with BSA, OxyBURST Green H 2 HFF BSA, as shown in Figure 1. Spectrofluorimetric analysis indicates that this reduced compound is non-fluorescent, and becomes fluorescent upon HRP-catalyzed oxidation.
Conversely, changes in lysosomal pH do not affect the fluorescence of oxidized OxyBURST Green H 2 HFF BSA as will be shown later.
The pH-dependency of fluorescence of both oxidized OxyBURST Green H 2 HFF BSA and 5-(and -6)-carboxyfluorescein were thus examined by the dual excitation ratiometric measurement in vitro , as shown in Figure 8. While 5-(and -6)-carboxyfluorescein exhibits strong pH-dependency, the experiment clearly demonstrates that fluorescence of the oxidized OxyBURST Green H 2 HFF BSA is not affected by variation of the pH within a range between 4.5 to 7.5.
To study the effect of pH on the fluorescence of oxidized OxyBURST Green H 2 HFF BSA and 5-(and -6)-carboxyfluorescein, 78 μg/mL of the oxidized dye or 4.2 μM 5-(and -6)-carboxyfluorescein were prepared in MES (4-morpholineethanesulfonic acid) buffer solutions containing 5 mM NaCl, 115 mM KCl, 1.2 mM MgSO 4 and 25 mM MES (pHs from 4.5 to 7.5).
The fluorescent image of oxidized OxyBURST Green H 2 HFF BSA in the same cell was then recorded as shown in Figure 9c.
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