Files

Abstract

Questions such as biomolecular interactions, kinetics of enzymes, DNA conformational changes, cellular structures properties (among others) are addressed by characterizing the biological function of molecules. This function depends on their structure and their dynamic properties, and are usually studied with optical labels. A widely used technique for that purpose is fluorescence correlation spectroscopy (FCS) that adopts fluorophores as reporters of dynamic processes. However, photobleaching limits the observation time on the one hand, and saturation at high excitation intensities imposes constraints for the study of fast molecular processes on the other hand. Gold nanoparticles (NPs) are labels of particular interest to overcome these drawbacks due to their photostability, low cytotoxicity and their availability with biocompatible surface coatings. In addition, NPs have found widespread application in life sciences and medicine, such as labels for optical imaging, carriers for drug and gene delivery, targets for cancer cell imaging and phototherapy, and contrast agent for magnetic resonance imaging (MRI). These applications all require methods to characterize their behaviour in a biological environment. Medical use of NPs usually requires intravenous injection. Once NPs are introduced into the blood stream they become exposed to biomolecules in the plasma that form a so-called protein corona. Obviously, knowledge about this interaction is needed for a safe medical application of NPs. The uptake of NPs by cells depends mainly on this protein corona. It is thus very important to understand the structural and dynamic properties of the protein corona at the molecular level to judge the fate of NPs in the human body prior to clinical studies. We introduce Optical Coherence Correlation Spectroscopy (OCCS) which exploits the backscattered light of NPs illuminated by a broadband light source. Due to the measurement of a scattering signal, OCCS gives access to long time scales that are hardly accessible with FCS experiments. The signal acquisition is similar to Fourier-domain optical coherence microscopy (FDOCM) with a dark-field contrast. We present the theory of OCCS based on auto-correlation analysis. Numerical simulations were used to estimate the performance of our new spectroscopy method and to build an analytical model for the auto-correlation function for measuring the diffusion coefficient and concentration of NPs in solution. An application to the study of the protein corona on NPs is presented. The hydrodynamic radius of superparamagnetic iron oxide nanoparticles (SPIONs) with adsorbed bovine serum albumin (BSA) is measured under physiological conditions using OCCS.We also aimed at improving the detection of these NPs in highly scattering environments, for instance inside a cell. Photothermal optical lock-in OCCS (poli-OCCS) is proposed, which exploits the absorption properties of gold NPs to generate a specific signal due to the photothermal effect. Poli-OCCS is introduced and first proof-of-principle measurements with gold NPs are presented.

Details

Actions