Centrifugation-based assay for examining nanoparticle–lipid membrane binding and disruption

Abstract

Physical disruption of cellular membranes arising from interactions with engineered nanoparticles is an important, but poorly understood aspect of nanotoxicology and nanomedicine. Model cellular membranes (i.e. lipid bilayers) can be used to identify interaction mechanisms, and most studies have largely focused on lipid bilayers supported on solid planar or spherical substrates. While useful and informative, these systems do not accurately represent an intact cell membrane because they restrict the elastic motion of the bilayer and the capacity for mechanical changes.Free standing bilayers are preferred, but add complexity. Given the importance of nanoparticle-membrane interactions in nanotoxicology and nanomedicine, and the vast range in nanoparticle composition, size, shape, and surface functionalization, there is a need to develop techniques that can rapidly and inexpensively analyze the membranenanoparticle activity by using free standing or unsupported membranes.This work develops a centrifugation-based assay that can analyze the membrane-nanoparticle activity as a function of nanoparticle surface functionalization, membrane lipid composition, and monovalent salt concentration (NaCl). Free standing, unsupported vesicles were used to gain relevant information on elastic membrane deformation and vesicle destabilization due to nanoparticle binding. Silver nanoparticles were chosen due to their widespread biological applications and surface plasmon resonance (SPR) properties. UV-vis based centrifugation assay, coupled with cryo-TEM and DLS analysis, was proposed to screen nanoparticle-membrane interactions; silver nanoparticles binding ratio RSPR was calculated as a function of Ag nanoparticle coating and vesicle composition. Study showed that strong electrostatic attraction led to significant sedimentation, vesicle / membrane disruption and higher RSPR value; in contrast, systems that exhibited weak or no electrostatic attraction did not show significant sedimentation, membrane disruption or high RSPR value. The centrifugation assay provides a rapid and straightforward way to screen nanoparticlemembrane interactions.iv ACKNOWLEDGMENTS

Salt-Assisted Graphene Oxide Dispersive Solid Phase Microextraction for Sensitive Detection of Malachite Green and Crystal Violet by HPLC

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“…Mechanisms underlying the antimicrobial activity of nanomaterials include (i) disruption of bacterial cell membrane (Xi and Bothun, 2014), (ii) induction of oxidative stress by free radical formation (von Moos and Slaveykova, 2014), (iii) mutagenesis (Ahmad et al, 2012), (iv) protein and DNA damage (Li et al, 2013), (v) inhibition of DNA replication by binding to DNA (Li et al, 2013), and (vi) respiratory chain disruption (Choi et al, 2008). Figure 3 depicts the plausible mechanisms by which metal or metal oxide NPs exhibit toxicity against bacteria and bacterial biofilm.…”