We employed the Monte Carlo simulation methodology to emulate the diffusion of fluorescently labeled particles and understand the source of differences between values of diffusion coefficients (and consequently hydrodynamic radii) of fluorescently labeled nanoparticles measured by fluorescence correlation spectroscopy (FCS) and dynamic light scattering (DLS). We used the simulation program developed in our laboratory and studied the diffusion of spherical particles of different sizes, which are labeled on their surface. In this study, we focused on two complicating effects: (i) multiple labeling and (ii) rotational diffusion which affect the fluorescence signal from large particles and hinder the analysis of autocorrelation functions according to simple analytical models. We have shown that the fluorescence fluctuations can be well fitted using the analytical model for small point-like particles, but the obtained parameters deviate in some cases significantly from the real ones. It means that the current data treatment yields apparent values of diffusion coefficients and other parameters only and the interpretation of experimental results for systems of particles with sizes comparable to the size of the active illuminated volume requires great care and precaution.
Determination of the tumor tissue optical properties during and after photodynamic therapy using inverse Monte Carlo method and double integrating sphere between 350 and 1000 nm
