The utility of asymmetric flow field-flow fractionation for preclinical characterization of nanomedicines
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Dynamic light scattering (DLS), transmission electron microscopy (TEM), and reversed phase-high performance liquid chromatography (RP-HPLC) are staples of nanoparticle characterization for size distribution, shape/morphology, and composition, respectively. These techniques are simple and provide important details on sample characteristics. However, DLS and TEM are routinely done in batch-mode, while RP-HPLC affords separation of components within the entire sample population, regardless of sample polydispersity. While batch-mode analysis is informative and should be a first-step analysis for any material, it may not be ideal for polydisperse formulations, such as many nanomedicines. Herein, we describe the utility of asymmetric flow field-flow fractionation (AF4) as a useful tool for a more thorough understanding of these inherently polydisperse materials. AF4 was coupled with in-line DLS for an enhanced separation and resolution of various size populations in a nanomaterial. Additionally, the various size populations were collected for offline analysis by TEM for an assessment of different shape populations, or RP-HPLC to provide a compositional analysis of each individual size population. This technique was also extended to assess nanoparticle stability, i.e., drug release, both in buffer and physiologically relevant matrix, as well as qualitatively evaluate the protein binding capacity of nanomedicines. Overall, AF4 is proven to be a very versatile technique and can provide a wealth of information on a material’s polydispersity and stability. Moreover, the ability to conduct analysis in physiological matrices provides an advantage that many other routine analytical techniques do not.
KeywordsNanoparticles Size distribution Polydispersity Protein binding Stability Drug release
The authors are grateful to Intezyne Technologies, Inc., Siva Therapeutics, and Cureport, Inc., for generously providing samples. The authors thank Joseph Meyer, Leidos Biomedical Research, Inc., for graphic illustrations.
This project has been funded in whole or in part with federal funds from the National Cancer Institute, National Institutes of Health, under Contract No. HHSN261200800001E.
Compliance with ethical standards
For plasma incubation studies, human plasma was collected from healthy volunteer donors under National Cancer Institute (NCI) at Frederick Protocol OH99-C-N046.
Conflict of interest
The authors declare that they have no competing interests.
The content of this publication does not necessarily reflect the views or policies of the Department of Health and Human Services, nor does mention of trade names, commercial products, or organizations imply endorsement by the US Government.
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