RESUMO
The encapsulation of therapeutic agents, such as drugs and vaccines, into colloidal particles offers an attractive strategy to enhance their efficacy. Previously, we reported the development of guanosine-based supramolecular colloidal particles suitable for encapsulating a broad array of guests ranging from small molecule drugs, like doxorubicin, to proteins, like GFP. Many biomedical applications of such particles require a precise determination of the amount of encapsulated therapeutic agents. Despite many studies describing the development of particle-based delivery systems, a general method for the precise and quick quantification of the encapsulated payload is still lacking. Here, we report a method based on flow cytometry measurements for complexes made from guanosine-based particles and a variety of commercially available fluorescent dyes. This method allows us to determine the apparent affinities of such dyes for two variants of these particles, which in turn provides insightful structure-affinity relationships. In contrast to the current methods, such as those that rely on fluorescence microscopy based on measurements of absorption/fluorescence of dissolved particles or on the supernatant of the solution, the reported method is suitable for high-throughput screening and more reproducible results. The protocol described here should be applicable to a wide variety of colloidal particles being developed around the world. Our group is currently expanding the scope to quantify the encapsulation of other molecules of biomedical interest, such as proteins and nucleic acids.
