LHA585

Guide (The following is our recommended protocol. This protocol is for guidance only and should be modified according to your specific needs).

1. Stock Solution Preparation
1.1 Solvent: Phosphate buffer (50 mM, pH = 7.4) containing 10% MeCN.
1.2 Concentration recommendation: 5 μM.

3. Staining Procedure
3.1 Sample type:[1]
3.1.1 Adherent cells (HeLa cells, HT22 cells).
3.2 Incubation conditions: Incubate with 5 μM LHA585 for 10 min; excitation wavelength 561 nm, emission wavelength 570-620 nm.
3.3 Washing steps: No washing required (wash-free imaging).

4. Controls
4.1 Positive controls: Cells treated with ferroptosis inducers including erastin (10 μM, 8 h), FIN56 (5 μM, 10 h), RSL3 (2 μM, 8 h).
4.2 Negative control: Cells pretreated with Fer-1 (2 μM, 8 h) followed by erastin (10 μM, 8 h).
4.3 Blank control: Untreated cells.

5. Detection & Analysis
5.1 Instrument: Confocal microscope.
5.1.1 Ex = 561 nm, Em = 570-620 nm.
5.2 Result analysis:
5.2.1 Fluorescence intensity changes: Significant fluorescence turn-on (ca. 360-fold in solution) in ferroptosis-induced cells; fluorescence intensity increases dose-dependently with ferroptosis inducer concentration; fluorescence signal rises earlier (within reperfusion initiation) compared to gold-standard probes in OGD/R models.
5.2.2 Fluorescence localization: Generated fluorescent rhodamine dye localizes to mitochondria.
5.2.3 Color changes: Red fluorescence emission.

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.