Diffusion coefficient of the fluorescent dye CF (carboxyfluorescein) in aqueous solution

Value 487 µm^2/sec Range: ±22 µm^2/sec
Organism Generic
Reference Kramer EM, Frazer NL, Baskin TI. Measurement of diffusion within the cell wall in living roots of Arabidopsis thaliana. J Exp Bot. 2007 58(11):3005-15PubMed ID17728296
Method To quantify the diffusion constant of small molecules in the plant cell wall, fluorescence from carboxyfluorescein (CF) in the intact roots of Arabidopsis thaliana was recorded. Roots were immersed in a solution of the fluorescent dye and viewed through a confocal fluorescence microscope. These roots are sufficiently transparent that much of the apoplast can be imaged. The diffusion coefficient, D(cw), of CF in the cell wall was probed using two protocols: fluorescence recovery after photobleaching and fluorescence loss following perfusion with dye-free solution. Diffusion coefficients were obtained from the kinetics of the fluorescent transients and modelling apoplast geometry. To determine the aqueous diffusion coefficient of CF, a small volume of the aqueous CF solution was placed between two coverslips, forming a thin film between 20 µm and 100 µm thick. Under the confocal microscope, a circular spot with a radius of ~200 µm and an optical thickness greater than the fluid thickness (to avoid measurement artefacts due to diffusion in the z-direction) was bleached. Then the half-time for fluorescence recovery, t1/2, was determined as the time needed for the intensity at the centre of the disc to reach 50% of the asymptotic recovery value. The exact solution to the diffusion equation, assuming an initial bleached disc of radius R, gives D=R^2/[4ln(2)t1/2]=(0.36R^2)/t1/2 (Crank, 1985). The aqueous diffusion coefficient for CF determined in this way was D=(4.87±0.22)×10^-10 m^2 s^-1 (mean±SE, n=4).
Comments See BNID 105033,105034
Entered by Uri M
ID 105032