Purification of DiD-Encapsulated Nanoparticles Through Triton-Washing

Researcher(s)

  • Emma Guzzetti, Biomedical Engineering, University of Delaware

Faculty Mentor(s)

  • Emily Day, Biomedical Engineering, University of Delaware

Abstract

The main goal of nanomedicine is to treat illnesses and diseases not only directly at their source but without damaging healthy tissues and functions or losing potency along the way (1). Therefore, it is imperative to understand exactly where nanoparticles (NPs) are ending up in the body. To study nanoparticle biodistribution, researchers track the location of encapsulated fluorophores. Dioctadecyl-3,3,3,3-tetramethylindodicarbocyanine (DiD) is part of a class of lipophilic carbocyanine dyes that are commonly used to track NP location (2-4). However, preliminary studies from our lab show this dye can form micelles of similar size and charge to the nanoparticles themselves, giving the appearance of successful encapsulation in NPs despite containing no polymeric material. This can lead to the dye giving a “false positive” of NP encapsulation during characterization and trials, leading to inaccurate results. The purpose of this study was to determine if micelles could be removed from DiD NPs using Triton X-100 (TX), an amphiphilic detergent. DiD-encapsulating Poly(lactic-co-glycolic acid)-Poly(ethylene-glycol) (PLGA-PEG) NPs and relevant controls were synthesized by nanoprecipitation and characterized by size, charge, and fluorescence. These NPs were then used to dose macrophages for one hour and 24 hours. Flow cytometry was used to quantify DiD signals at each timepoint. The non-TX DiD NPs were smaller but fluoresced more compared to the TX DiD NPs. The smaller size and higher fluorescence can be attributed to the extra dye in the solution affecting the results. Future research should continue to study how to remove the micelles, allowing for more accurate NP location testing and overall helping nanomedicine produce safer and more effective therapeutics.

References

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