Liperfluo is an analog of Spy‑LHP and can be used for the detection of lipid peroxides (LPO). Liperfluo is specifically oxidized by lipid peroxides and emits strong fluorescence in organic solvents such as ethanol (Figures 1, 2). The oxidized Liperfluo exhibits excitation and emission wavelengths at 524 nm and 535 nm, respectively (both long‑wavelength), which reduces photodamage to test samples and decreases sample autofluorescence. Bearing a tetraethylene glycol moiety attached to one terminus of the diisoquinoline ring, Liperfluo possesses improved molecular dispersibility in aqueous solution. Although its oxidized form shows barely any fluorescence in water, it generates intense fluorescence in lipid‑rich compartments such as cell membranes. Accordingly, Liperfluo is suitable for fluorescence imaging or flow cytometric analysis of lipid peroxides in living cells.
Features
1. Enables imaging and detection of lipid peroxides in cells;
2. Long‑wavelength excitation mitigates cellular photodamage and interference from autofluorescence;
3. High specificity toward lipid peroxides.
Research Areas
Ferroptosis research: Liperfluo is applied in the study of ferroptosis, one form of regulated cell death.
Troubleshooting & Solution
1. Preparation of 1 mmol/L Liperfluo DMSO Solution
· Add 60 μL DMSO to the Liperfluo vial, and pipette up‑and‑down 30 times within 1 min. <sup>*1,*2,*3</sup>
· Wrap the vial with aluminum foil to protect from light, then vortex for 5 min.
Once dissolved, the Liperfluo‑DMSO solution shall be stored protected from light and used within 1 day.
*1 Liperfluo is light‑sensitive; perform pipetting within 1 minute.
*2 If solid Liperfluo adheres to the bottom of the vial, crush large solid chunks into small pieces with the pipette tip during pipetting.
*3 If solids remain after vortexing, sonicate or heat the vial (40 °C, 30 min) to achieve full dissolution. Ensure the lower portion of the vial is submerged in water during sonication. See the figure below for details.
2. Preparation of 1 mmol/L Liperfluo DMSO Stock Solution
The optimal Liperfluo concentration for staining varies with experimental conditions. Follow the procedures below for fluorescence microscopy:
Fluorescence imaging: Dilute the 1 mmol/L Liperfluo DMSO stock with serum‑free medium to prepare the working solution.
Flow cytometry: Add the 1 mmol/L Liperfluo DMSO stock directly into the cell suspension to be stained.
Operating Procedure
Detection of lipid peroxides in cells
1. Seed cells in culture dishes or well plates.
2. Discard supernatant and wash cells once with serum‑free medium.
3. Add an appropriate volume of Liperfluo working solution, and incubate at 37 °C for 30 min. *The optimal Liperfluo concentration depends on experimental conditions; condition optimization in advance is required.
4. Remove supernatant and wash cells twice with serum‑free medium.
5. Observe cells under a fluorescence microscope or analyze cells by flow cytometry.
Fluorescence spectra
