FIGURE 3.
FCS experiment of internally labeled spheres with radius A = 0.5 μm. (A) The intensity trace of the data taken at 2 kHz for 3 h. Each spike represents a sphere passing through the observation volume. (B and C) The experimental correlation function (○) and its fit (solid line) to the point particle model (B) and solid sphere model (C). The beam waist of the PSF is fixed to 0.42 μm, which is determined by calibration with a sample of 0.025-μm spheres, as described in the text. The fit to a point particle model yields a radius A = 1.53 ± 0.17 μm and a diffusion time τd = 238 ± 27 ms, which overestimates the size threefold. If the same data is fit to a solid sphere model, we obtain A = 0.48 ± 0.02 μm and a diffusion time τd = 75 ± 4 ms, which agrees very well with the prediction based on the Stokes-Einstein relation.