IMAGING OF FLUOROPHORES IN CHROMATOGRAPHIC BEADS, RECONSTRUCTION OF RADIAL DENSITY DISTRIBUTIONS AND CHARACTERISATION OF PROTEIN UPTAKING PROCESSES

Abstract

A new adjustment calculus is presented to determine the true intraparticle distribution of bound protein within chromatographic beads from confocal fluorescence slice series. The calculus does not require knowledge about optical properties of different chromatographic materials like refractive index and turbidity, but it depends on a parameter which can be adjusted interactively. The algorithm is of complexity O(n) where n is the pixel number. From the reconstructed data we compute the parameters of the protein uptaking process using a model-based approach. It is demonstrated that the protein uptaking rates of the beads strongly dependent on the conditions of the fluid phase influencing the strength of protein surface interaction.

Keywords

• bead characterisation
• chromatography
• confocal laser scanning microscopy
• image reconstruction
• light attenuation