Optics-intrinsic double-circle phenomenon in protein adsorption visualized by confocal laser scanning microscopy

Confocal laser scanning microscopy (CLSM) has been introduced in the research of chromatographic adsorption and the technique provides a direct and noninvasive way to measure and characterize intraparticle concentration distributions of proteins on a single particle level. In this work, an optics-intrinsic double-circle phenomenon that occurs in protein adsorption visualized by CLSM is exhibited and its optical mechanism is reasonably explained by an evolutive version of the attenuation equation. The time course of intraparticle fluorescence intensity profiles obtained from CLSM (even the double-circle phenomenon) in this research is vividly reproduced by model simulation, which takes into account the light attenuation by combining the attenuation equation with an effective pore diffusion model.