We have used the technique of fluorescence photobleaching recovery to measure the lateral diffusion coefficients and the mobile fractions of a fluorescent lipid probe, 1-acyl-2-(12-[(7-nitro-2-1,3-benzoxadiazol-4-yl)aminododecanoyl]) phosphatidylcholine (NBD-PC), and of labeled membrane proteins of human fibroblasts. Values for mobile fractions decrease monotonically with increasing size of the laser spot used for the measurements, over a range of 0.35-5.0 μm. Values for NBD-PC diffusion coefficients increase in part of this range to reach a plateau at larger laser spots. This variation is not an artifact of the measuring system, since the effects are not seen if diffusion of the probe is measured in liposomes. We also find that the distribution of diffusion coefficients measured with small laser spots is heterogeneous indicating that these small spots can sample different regions of the membrane. These regions appear to differ in protein concentration. Our data strongly indicate that fibroblast surface membranes consist of protein-rich domains ~1 μm in diameter, embedded in a relatively protein-poor lipid continuum. These features appear in photographs of labeled cell surfaces illuminated by the expanded laser beam.
ASJC Scopus subject areas
- Cell Biology