A high throughput method for quantification of cell surface bound and internalized chitosan nanoparticles

Chitosan has become a popular polymer for drug delivery. It's hydro solubility and mild formulation conditions have made it an attractive polymer for macromolecular delivery. Accurate quantification of internalized chitosan nanoparticles (NPs) is imperative for fair assessment of the nano-formulation where it is important to determine the exact amount of drug actually being delivered into the cell, especially for macromolecular drugs where cellular entry is limited by molecule size and/or charge. The preferential affinity of wheat germ agglutinin tagged with fluorescein isothiocyanate (WGA-FITC) to chitosan is exploited in the development of a simple and rapid method for the differentiation between and quantification of cell surface bound and internalized chitosan NPs. The percentage of cell surface bound NPs could be easily determined and corrected NP uptake could be calculated accordingly. The developed method is applicable in several cell lines and has successfully been tested with NPs with different sizes (25 and 150 nm) and with very low NP concentrations (20 mu g/mL). The method will allow for the correct evaluation of chitosan NP uptake and could be further used to evaluate chitosan based nanomedicine and provide guidelines on how to modify NPs for enhanced internalization, and improved drug delivery. (C) 2015 Elsevier B.V. All rights reserved.