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The protein corona, which forms on the nanoparticle's surface in most biological media, determines the nanoparticles physicochemical characteristics. The formation of the protein corona has a significant impact on the biodistribution and clearance of nanoparticles in vivo. Therefore, the ability to influence the formation of the protein corona is essential to most biomedical applications, including drug delivery and imaging. In this study, we investigate the protein adsorption on nanoparticles with a hydrodynamic radius of 30 nm and a coating of thermoresponsive poly(2-isopropyl-2-oxazoline) in serum. Using multiangle dynamic light scattering (DLS) we demonstrate that heating of the nanoparticles above their phase separation temperature induces the formation of agglomerates, with a hydrodynamic radius of 1 µm. In serum, noticeably stronger agglomeration occurs at lower temperatures compared to serum-free conditions. Cryogenic transmission electron microscopy (cryo-TEM) revealed a high packing density of agglomerates when serum was not present. In contrast, in the presence of serum, agglomerated nanoparticles were loosely packed, indicating that proteins are intercalated between them. Moreover, an increase in protein content is observed upon heating, confirming that protein adsorption is induced by the alteration of the surface during phase separation. After cooling and switching the surface back, most of the agglomerates were dissolved and the main fraction returned to the original size of approximately 30 nm as shown by asymmetrical flow-field flow fractionation (AF-FFF) and DLS. Furthermore, the amounts of adsorbed proteins are similar before and after heating the nanoparticles to above their phase-separation temperature. Overall, our results demonstrate that the thermoresponsivity of the polymer coating enables turning the corona formation on nanoparticles on and off in situ. As the local heating of body areas can be easily done in vivo, the thermoresponsive coating could potentially be used to induce the agglomeration of nanoparticles and proteins and the accumulation of nanoparticles in a targeted body region.
In biological fluids, proteins associate with nanoparticles, leading to a protein 'corona' defining the biological identity of the particle. However, a comprehensive knowledge of particle-guided protein fingerprints and their dependence on nanomaterial properties is incomplete. We studied the long-lived ('hard') blood plasma derived corona on monodispersed amorphous silica nanoparticles differing in size (20, 30, and 100 nm). Employing label-free liquid chromatography mass spectrometry, one- and two-dimensional gel electrophoresis, and immunoblotting the composition of the protein corona was analyzed not only qualitatively but also quantitatively. Detected proteins were bioinformatically classified according to their physicochemical and biological properties. Binding of the 125 identified proteins did not simply reflect their relative abundance in the plasma but revealed an enrichment of specific lipoproteins as well as proteins involved in coagulation and the complement pathway. In contrast, immunoglobulins and acute phase response proteins displayed a lower affinity for the particles. Protein decoration of the negatively charged particles did not correlate with protein size or charge, demonstrating that electrostatic effects alone are not the major driving force regulating the nanoparticle–protein interaction. Remarkably, even differences in particle size of only 10 nm significantly determined the nanoparticle corona, although no clear correlation with particle surface volume, protein size, or charge was evident. Particle size quantitatively influenced the particles decoration with 37% of all identified proteins, including (patho)biologically relevant candidates. We demonstrate the complexity of the plasma corona and its still unresolved physicochemical regulation, which need to be considered in nanobioscience in the future.
Due to the adsorption of biomolecules, the control of the biodistribution of nanoparticles is still one of the major challenges of nanomedicine. Poly(2-ethyl-2-oxazoline) (PEtOx) for surface modification of nanoparticles is applied and both protein adsorption and cellular uptake of PEtOxylated nanoparticles versus nanoparticles coated with poly(ethylene glycol) (PEG) and non-coated positively and negatively charged nanoparticles are compared. Therefore, fluorescent poly(organosiloxane) nanoparticles of 15 nm radius are synthesized, which are used as a scaffold for surface modification in a grafting onto approach.
With multi-angle dynamic light scattering, asymmetrical flow field-flow fractionation, gel electrophoresis, and liquid chromatography-mass spectrometry, it is demonstrated that protein adsorption on PEtOxylated nanoparticles is extremely low, similar as on PEGylated nanoparticles. Moreover, quantitative microscopy reveals that PEtOxylation significantly reduces the non-specific cellular uptake, particularly by macrophage-like cells. Collectively, studies demonstrate that PEtOx is a very effective alternative to PEG for stealth modification of the surface of nanoparticles.