Laurdan

Laurdan is a membrane-permeable fluorescent probe that displays spectral sensitivity to the phospholipid phase of the cell membrane to which it is bound. Quantitation of generalized polarization (GP) of Laurdan can be used to identify phospholipid phase (Ex/Em = 370/440-490 nm). In Vitro: Laurdan (from 1:1000 to 5:1000) acts as a biosensor to monitor changes in the membrane water content during cell migration in NIH3T3 cells^[1].
Laurdan staining solution preparation
1.1 Preparation of stock solution
Dissolve Laurdan in dimethyl sulfoxide (DMSO) to prepare a 1.8 mM stock solution.
1.2 Preparation of working solution
Dilute the Laurdan stock solution at a dilution of 1:1000.
Laurdan staining
1) NIH3T3 cells were cultured in Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum, 5 mL Pen-Strep, and 2.5 mL 1 M HEPES.
2) Freshly isolated NIH3T3 cells were seeded into 35 mm Mattek glass-bottom dishes coated with fibronectin.
3) Cells were incubated with Laurdan for 40 min at 37°C.
4) Observed using an LSM710 META laser scanning microscope.
5) SimFCS software processing to calibrate the system and phasor diagram of FLIM data by measuring fluorescein (pH 11).