Membrane structure in which proteins and lipids have relative lateral mobility; the cell membrane is composed of a phospholipid bilayer and proteins that are embedded, threaded through it, and adsorbed on its surface, with the hydrophobic tail of the phospholipid bilayer inside and the hydrophilic head outside.

Operation method

fluorescent probe labeling method

Principle

A fluorescent probe commonly used to study membrane lipid fluidity is 1,6-diphenyl-1,3,5-hexatriene (DPH).After DPH is doped into the hydrocarbon chain region of cellular membrane lipids, the viscosity of the medium is increased and cis-trans isomerism is inhibited, making it the only all-trans conformation capable of luminescence.

Materials and Instruments

Tumor Cells
Phosphate Buffer DPH
Fluorescence Spectrophotometer Centrifuge

Move

1. Tumor cells in logarithmic growth phase were taken and washed twice with pH 7.2, 0.01 mol/l phosphate buffer.

2. ¹⁰⁷ cells were taken, centrifuged, and 2 ml of ^2×106 mol/l DPH solution was added.

3. incubate at 25°C with oscillation for 30 min, and then wash once with PBS.

4. Suspend in 4 ml PBS solution.

5. The measuring instrument was a fluorescence spectrophotometer, and the fluorescence intensity was measured at the excitation wavelength of 362 nm, emission wavelength of 432 nm and slit 10.

Caveat

Since DPH can enter the cell interior, its cationic ionic derivative 1-(4-trimethylaminophenyl)-6-phenyl-1,3,5-hexatriene (TMDPH) was later synthesized.TMDPH could not enter the cell through the cell membrane, so theoretically, TMDPH labeling method can more accurately reflect the membrane lipid mobility than DPH labeling method.

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