Fluorescence correlation spectroscopy in an optical trap

Fluorescence correlation spectroscopy is one of the most sensitive methods for biomolecule and nanoparticle studies. By fluorescently labeling the target particles, statistical analysis of the fluorescence fluctuation inside the focal volume of a laser beam yield information of the concentration and diffusion of the target particles in the illuminated volume. However, the application of FCS is limited by its detection range of 1010-1014 particles/mL. To overcome this sensitivity threshold on the low concentration end, we designed a hybrid system that augments FCS with optical trapping. By using the optical gradient force from a second laser focused to the same illuminated volume, we were able to show that the local concentration of particles can be enriched significantly, thus extending the useful range of FCS. In this work, we describe this novel hybrid optical method for nanoparticle detection by first considering freely diffusing particles about the illuminating volume, and then compare the results to nanoparticles under the influence of the optical trapping laser. Analysis of trapped particle number permits measurement of trapping energy as well as determine ambient concentration out of the trap. Furthermore, the hindered diffusion of trapped particles due to optical forces will be discussed.