Winzen S, Schoettler S, Baier G, Rosenauer C, Mailaender V, Landfester K, Mohr K
Max Planck Institute of Polymer Research, Ackermannweg 10, 55128 Mainz, Germany.
Nanoscale. 2015 Feb 21;7(7):2992-3001. doi: 10.1039/c4nr05982d.
Here we demonstrate how a complementary analysis of nanocapsule-protein interactions with and without application media allows gaining insights into the so called hard and soft protein corona. We have investigated how both human plasma and individual proteins (human serum albumin (HSA), apolipoprotein A-I (ApoA-I)) adsorb and interact with hydroxyethyl starch (HES) nanocapsules possessing different functionalities. To analyse the hard protein corona we used sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and a protein quantitation assay. No significant differences were observed with regards to the hard protein corona. For analysis of the soft protein corona we characterized the nanocapsule-protein interaction with isothermal titration calorimetry (ITC) and dynamic light scattering (DLS). DLS and ITC measurements revealed that a high amount of plasma proteins were adsorbed onto the capsules' surface. Although HSA was not detected in the hard protein corona, ITC measurements indicated the adsorption of an HSA amount similar to plasma with a low binding affinity and reaction heat. In contrast, only small amounts of ApoA-I protein adsorb to the capsules with high binding affinities. Through a comparison of these methods we have identified ApoA-I to be a component of the hard protein corona and HSA as a component of the soft corona. We demonstrate a pronounced difference in the protein corona observed depending on the type of characterization technique applied. As the biological identity of a particle is given by the protein corona it is crucial to use complementary characterization techniques to analyse different aspects of the protein corona.
在这里，我们展示了对有无应用介质时纳米胶囊 - 蛋白质相互作用进行补充分析，如何有助于深入了解所谓的硬蛋白冠层和软蛋白冠层。我们研究了人血浆以及单个蛋白质（人血清白蛋白（HSA）、载脂蛋白A - I（ApoA - I））如何吸附到具有不同功能的羟乙基淀粉（HES）纳米胶囊上并与其相互作用。为了分析硬蛋白冠层，我们使用了十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS - PAGE）和蛋白质定量测定法。在硬蛋白冠层方面未观察到显著差异。对于软蛋白冠层的分析，我们用等温滴定量热法（ITC）和动态光散射（DLS）对纳米胶囊 - 蛋白质相互作用进行了表征。DLS和ITC测量结果表明，大量血浆蛋白吸附到了胶囊表面。虽然在硬蛋白冠层中未检测到HSA，但ITC测量表明有与血浆中含量相似的HSA以低结合亲和力和反应热被吸附。相比之下，只有少量ApoA - I蛋白以高结合亲和力吸附到胶囊上。通过这些方法的比较，我们确定ApoA - I是硬蛋白冠层的一个组分，而HSA是软蛋白冠层的一个组分。我们证明，根据所应用的表征技术类型，观察到的蛋白冠层存在显著差异。由于颗粒的生物学特性由蛋白冠层决定，因此使用互补的表征技术来分析蛋白冠层的不同方面至关重要。
