Andreas Henkel

Single Unlabeled Protein Detection on Individual Plasmonic Nanoparticles

The ultimate detection limit in analytic chemistry and biology is the single molecule. Commonly, fluorescent dye labels or enzymatic amplification are employed. This requires additional labeling of the analyte, which modifies the species under investigation and therefore influences biological processes. Here, we utilize single gold nanoparticles to detect single unlabeled proteins with extremely high temporal resolution. This allows for monitoring the dynamic evolution of a single protein binding event on a millisecond time scale. The technique even resolves equilibrium coverage fluctuations, opening a window into Brownian dynamics of unlabeled macromolecules. Therefore, our method enables the study of protein folding dynamics, protein adsorption processes, and kinetics as well as nonequilibrium soft matter dynamics on the single molecule level.

Multiplexed Plasmon Sensor for Rapid Label-Free Analyte Detection

Efficient and cost-effective multiplexed detection schemes for proteins in small liquid samples would bring drastic advances to fields like disease detection or water quality monitoring. We present a novel multiplexed sensor with randomly deposited aptamer functionalized gold nanorods. The spectral position of plasmon resonances of individual nanorods, monitored by dark-field spectroscopy, respond specifically to different proteins. We demonstrate nanomolar sensitivity, sensor recycling, and the potential to upscale to hundreds or thousands of targets.

A new approach to assess gold nanoparticle uptake by mammalian cells: Combining optical dark-field and transmission electron microscopy

Toxicological effects of nanoparticles are associated with their internalization into cells. Hence, there is a strong need for techniques revealing the interaction between particles and cells as well as quantifying the uptake at the same time. For that reason, herein optical dark-field microscopy is used in conjunction with transmission electron microscopy to investigate the uptake of gold nanoparticles into epithelial cells with respect to shape, stabilizing agent, and surface charge. The number of internalized particles is strongly dependent on the stabilizing agent, but not on the particle shape. A test of metabolic activity shows no direct correlation with the number of internalized particles. Therefore, particle properties besides coating and shape are suspected to contribute to the observed toxicity.

Plasmonic nanosensors for simultaneous quantification of multiple protein-protein binding affinities.

Most of current techniques used for the quantification of protein-protein interactions require the analysis of one pair of binding partners at a time. Herein we present a label-free, simple, fast, and cost-effective route to characterize binding affinities between multiple macromolecular partners simultaneously, using optical dark-field spectroscopy and individual protein-functionalized gold nanorods as sensing elements. Our NanoSPR method could easily become a simple and standard tool in biological, biochemical, and medical laboratories.

Surface Asymmetry of Coated Spherical Nanoparticles

We validate the nonspherical grafting arrangement of isotropically coated spherical nanoparticles as very recently proposed. We utilize localized surface plasmon resonance enhanced dynamic polarized and depolarized light scattering from Au nanoparticles, the spherical symmetry of which was revealed by single-particle dark-field spectroscopy. The same Au nanospheres are grafted with ligands of different chemistry and length. The wavelength dependent depolarization ratio and the two transport coefficients of these nanoparticles, obtained from the dynamic light scattering experiment, can only be reconciled with the TEM data, the single UV/vis extinction spectrum, and the dark-field spectroscopy experiments if their coating is described as asymmetric. Spatially anisotropic graft distribution on spherical nanoparticles impacts their assembly and understanding its origin will help control the structure and properties of polymer nanocomposites.

Single Particle Plasmon Sensors as Label-Free Technique To Monitor MinDE Protein Wave Propagation on Membranes

We use individual gold nanorods as pointlike detectors for the intrinsic dynamics of an oscillating biological system. We chose the pattern forming MinDE protein system from Escherichia coli (E. coli), a prominent example for self-organized chemical oscillations of membrane-associated proteins that are involved in the bacterial cell division process. Similar to surface plasmon resonance (SPR), the gold nanorods report changes in their protein surface coverage without the need for fluorescence labeling, a technique we refer to as NanoSPR. Comparing the dynamics for fluorescence labeled and unlabeled proteins, we find a reduction of the oscillation period by about 20%. The absence of photobleaching allows us to investigate Min proteins attaching and detaching from lipid coated gold nanorods with an unprecedented bandwidth of 100 ms time resolution and 1 h observation time. The long observation reveals small changes of the oscillation period over time. Averaging many cycles yields the precise wave profile that exhibits the four phases suggested in previous reports. Unexpected from previous fluorescence-based studies, we found an immobile static protein layer not dissociating during the oscillation cycle. Hence, NanoSPR is an attractive label-free real-time technique for the local investigation of molecular dynamics with high observation bandwidth. It gives access to systems, which cannot be fluorescently labeled, and resolves local dynamics that would average out over the sensor area used in conventional SPR.

Silica-coated Au@ZnO Janus particles and their stability in epithelial cells

Multicomponent particles have emerged in recent years as new compartmentalized colloids with two sides of different chemistry or polarity that have opened up a wide field of unique applications in medicine, biochemistry, optics, physics and chemistry. A drawback of particles containing a ZnO hemisphere is their low stability in biological environment due to the amphoteric properties of Zn2+. Therefore we have synthesized monodisperse Au@ZnO Janus particles by seed-mediated nucleation and growth whose ZnO domain was coated selectively with a thin SiO2 layer as a protection from the surrounding environment that imparts stability in aqueous media while the Au domain remained untouched. The thickness of the SiO2 layer could be precisely controlled. The SiO2 coating of the oxide domain allows biomolecule conjugation (e.g. antibodies, proteins) in a single step for converting the photoluminescent and photocatalytic active Janus nanoparticles into multifunctional efficient vehicles for cell targeting. The SiO2-coated functionalized nanoparticles were stable in buffer solutions and other aqueous systems. Biocompatibility and potential biomedical applications of the Au@ZnO@SiO2 Janus particles were assayed by a cell viability analysis by co-incubating the Au@ZnO@SiO2 Janus particles with epithelia cells and compared to those of uncoated ZnO.

Synthesis of Au–CdS@CdSe Hybrid Nanoparticles with a Highly Reactive Gold Domain

We propose a novel route to synthesize semiconductor-gold hybrid nanoparticles directly in water, resulting in much larger gold domains than previous protocols (up to 50 nm) with very reactive surfaces which allow further functionalization. This method advances the possibility of self-assembly into complex structures with catalytic activity toward the reduction of nitro compounds by hydrides. The large size of these gold domains in hybrid particles supports efficient light scattering at the plasmon resonance frequency, making such structures attractive for single-particle studies.

En-Face differential absorption optical coherence tomography with gold nanorods as the contrast agent

A new variety of nanoparticles showing unique and characteristic optical properties, appeals for its use as contrast agents in medical imaging. Gold nanospheres, nanorods and nanoshells with a silica core are new forms of promising contrast agents which can be tuned to specific absorption or scattering characteristics within the near-infrared (NIR) spectrum ranging from 650 - 1300 nm. They have the ability to be used for both image enhancement and as photosensitive markers due to their well designable scattering and absorption properties. Furthermore, their strong optical absorption permits treatment of malignant cells by photoablation processes, induced when heating them with a matched light source. Differential absorption optical coherence tomography (DA-OCT) allows for the detection and depth resolved concentration measurement of such markers. So far, reports on DA-OCT systems used A-scan based imaging systems to assess depth resolved information about the absorption properties and the concentration of a chemical compound. Enface OCT (B(T) or C(T) scan based) images allow for better depth localization and a depth resolved concentration measurement of the compound under investigation. For this aim, we evaluate the suitability of a multiscan time-domain OCT set-up, compatible with different light sources providing different wavelengths and bandwidths in the NIR, to perform differential absorption OCT measurements, using gold nanorods as the contrast agent.