Accurate assessment of nanomedicines' antibacterial properties is pivotal for their effective use in both in vitro and in vivo settings. Conventional antibacterial activity assessment methods, involving bacterial coculture with compounds on agar plates, may not fully suit nanomedicines due to their susceptibility to alterations in physicochemical properties induced by biological fluids. Furthermore, these biological fluids might even enhance the bacterial growth. This study introduces a novel, rigorous, and reproducible methodology for evaluating nanomedicine antibacterial properties using cell culture media (i.e., DMEM-FBS10%). To assess the antibacterial activity of the nanoparticles in cell culture media, superparamagnetic iron oxide nanoparticles (SPIONs) were chosen as the model nanomedicine due to their clinical significance. A comparative analysis between the traditional and our proposed methods yielded contrasting outcomes, shedding light on the significant impact of biological fluids on nanoparticle antibacterial activities. While the conventional approach suggested the antibacterial effectiveness of SPIONs against Staphylococcus aureus, our innovative method unveiled a substantial increase in bacterial growth in the presence of biological fluids. More specifically, we found a significant increase in bacterial growth when exposed to bare SPIONs at various concentrations, while the formation of a protein corona on SPION surfaces could markedly reduce the observed bacterial growth compared to the control group. These findings underscore the necessity for more refined evaluation techniques that can better replicate the in vivo environment when studying the nanomedicine's antibacterial capabilities.
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