Laurdan

Biological Activity

Laurdan is a membrane-permeable fluorescent probe that is spectrally sensitive to the phospholipid phase of cell membranes to which it binds, and can be used for the identification of phospholipid phases, the absorption wavelength of Laurdan is 340-380nm and the emission wavelength is 440-490nm. Laurdan is phase-sensitive. Its fluorescence emission in the liquid crystal phase will show a large red shift, while it will show a small red shift in the gel phase, which can detect the phase state and fluidity of the membrane. Laurdan can measure the generalized polarization (GP) of the cell membrane. High GP is usually associated with low fluidity, low polarity or high cholesterol content of the membrane, while low GP is the opposite.

The following experimental protocol only provides a reference for a Laurdan generalized polarization (GP) measurement experiment of cell membrane. Please modify the working concentration and staining time according to experimental needs or refer to the literature. (from the literature, for reference only)

1. Solution preparation

(1) Storage solution: Dissolve Laurdan in DMSO or chloroform to a final concentration of 5 mM.
Note: After unused storage solution is aliquoted, store it at -20℃ in the dark to avoid repeated freezing and thawing.

(2) Working solution: Dilute the storage solution with experimental buffer (e.g. cell culture medium) to the required working concentration, usually in the range of 1-10 µM.
Note: Please adjust the optimal working concentration according to the actual situation or refer to the literature to set the gradient concentration by yourself. The working solution must be prepared and used immediately.

2. Steps for measuring the generalized polarization (GP) of Bacillus subtilis cell membrane by Laurdan

(1) Cell treatment: Cultivate cells to the middle of the logarithmic growth phase, wash them four times in PBS containing 2% glucose and 1% DMF, resuspend them in the same buffer, and adjust the OD600 to 0.3.

(2) Liposome preparation: Bacillus subtilis polar lipid extract was prepared by detergent dialysis, and the prepared liposome solution was squeezed through a 0.4μm filter membrane 20 times. The 10mg/ml liposome stock solution was diluted to a concentration of 1 mg/ml with Tris buffer (5mM, pH 7.4).

(3) Liposome labeling: Add Laurdan solution to the cell suspension to a final concentration of 10 μM and incubate in the dark for 30 minutes.

(4) Fluorescence measurement: Laurdan fluorescence measurement was performed using a BioTek Synergy MX plate reader. The excitation wavelength was 350 nm, the emission wavelengths were 460 nm and 500 nm, and readings were taken every 2 minutes.

(5) Laurdan fluorescence polarization calculation: Laurdan generalized polarization (GP) value was calculated using the following formula: (I460-I500)/(I460+I500).

References: https://pubmed.ncbi.nlm.nih.gov/29451901/

Chemical Information

Molecular Weight	353.54
Formula	C24H35NO
CAS Number	74515-25-6
Form	Solid
Solubility (25°C)	DMSO 5 mg/mL Chloroform 10 mg/mL
Storage	Powder 4°C, protect from light

References

[1] Hanna Orlikowska-Rzeznik et al. J Phys Chem B. Laurdan Discerns Lipid Membrane Hydration and Cholesterol Content

[2] Irena Levitan. Curr Top Membr. Evaluating membrane structure by Laurdan imaging: Disruption of lipid packing by oxidized lipids

[3] Michaela Wenzel, et al. Bio Protoc. Assessing Membrane Fluidity and Visualizing Fluid Membrane Domains in Bacteria Using Fluorescent Membrane Dyes

[4] German Gunther et al. Acc Chem Res. LAURDAN since Weber: The Quest for Visualizing Membrane Heterogeneity

[5] Dennapa Saeloh, et al. PLoS Pathog. The novel antibiotic rhodomyrtone traps membrane proteins in vesicles with increased fluidity

[6] Kathi Scheinpflug, et al. Methods Mol Biol. Measurement of Cell Membrane Fluidity by Laurdan GP: Fluorescence Spectroscopy and Microscopy

[7] Anthony G Jay et al. J Fluoresc. Disorder Amidst Membrane Order: Standardizing Laurdan Generalized Polarization and Membrane Fluidity Terms

[8] Silvia S Antollini et al. Methods Mol Biol. Laurdan studies of membrane lipid-nicotinic acetylcholine receptor protein interactions