Spy-LHP is a newly developed fluorescent probe for live cell imaging of phospholipid peroxide. There are several detection methods available for lipid peroxides, such as iodide titration method, colorimetric method, or chemiluminometric method to determine malondialdehyde or 4-hydroxynonenal. Malondialdehyde and 4-hydroxynonenal are derivatives from lipid hydroperoxide prepared by oxidation with reactive oxygen species. Thiobarbituriic acid and 1-Methyl-2-phenylindole are used for deriving malondialdehyde for the colorimetric or fluorometric analysis.
Spy-LHP is a low-fluorescent compound, but is oxidized with lipid hydroperoxide to become a high fluorescent compound as indicated in Fig. 1. A similar product, DPPP, is oxidized by a lipid hydroperoxide and becomes a fluorescence compound that can be excited at 352 nm to emit fluorescence at 380 nm. However, the UV excitation for DPPP significantly damages a live cell. Since the oxidized Spy-LHP emits
strong fluorescence (quantum yield: ~1) with maximum wavelength at 535 nm when excited at 524 nm, damage to live cells is very small. Spy-LHP has two alkyl chains to improve the affinity to the lipid bilayer. Spy-LHP is highly selective to lipid hydroperoxide and does not react with hydrogen peroxide, hydroxy radicals, superoxide anion, nitric oxides, peroxynitrite, and alkylperoxy radicals.
Fig. 1 Reaction of Spy-LHP with lipid hydroperoxide
1 mg/150µL acetone (25oC, saturated) or 1 mg/ml （chloroform)
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