Showing papers by "Peter Zijlstra published in 2020"

Plasmonic Assemblies for Real‐Time Single‐Molecule Biosensing

Abstract: Their tunable optical properties and versatile surface functionalization have sparked applications of plasmonic assemblies in the fields of biosensing, nonlinear optics, and photonics. Particularly, in the field of biosensing, rapid advances have occurred in the use of plasmonic assemblies for real-time single-molecule sensing. Compared to individual particles, the use of assemblies as sensors provides stronger signals, more control over the optical properties, and access to a broader range of timescales. In the past years, they have been used to directly reveal single-molecule interactions, mechanical properties, and conformational dynamics. This review summarizes the development of real-time single-molecule sensors built around plasmonic assemblies. First, a brief overview of their optical properties is given, and then recent applications are described. The current challenges in the field and suggestions to overcome those challenges are discussed in detail. Their stability, specificity, and sensitivity as sensors provide a complementary approach to other single-molecule techniques like force spectroscopy and single-molecule fluorescence. In future applications, the impact in real-time sensing on ultralong timescales (hours) and ultrashort timescales (sub-millisecond), time windows that are difficult to access using other techniques, is particularly foreseen.

Continuous Small-Molecule Monitoring with a Digital Single-Particle Switch.

Abstract: The ability to continuously measure concentrations of small molecules is important for biomedical, environmental, and industrial monitoring. However, because of their low molecular mass, it is difficult to quantify concentrations of such molecules, particularly at low concentrations. Here, we describe a small-molecule sensor that is generalizable, sensitive, specific, reversible, and suited for continuous monitoring over long durations. The sensor consists of particles attached to a sensing surface via a double-stranded DNA tether. The particles transiently bind to the sensing surface via single-molecular affinity interactions, and the transient binding is optically detected as digital binding events via the Brownian motion of the particles. The rate of binding events decreases with increasing analyte concentration because analyte molecules inhibit binding of the tethered particle to the surface. The sensor enables continuous measurements of analyte concentrations because of the reversibility of the intermolecular bonds and digital read-out of particle motion. We show results for the monitoring of short single-stranded DNA sequences and creatinine, a small-molecule biomarker (113 Da) for kidney function, demonstrating a temporal resolution of a few minutes. The precision of the sensor is determined by the statistics of the digital switching events, which means that the precision is tunable by the number of particles and the measurement time.

Dynamic single-molecule counting for the quantification and optimization of nanoparticle functionalization protocols

Abstract: Applications of colloidal particles in the fields of i.e. biosensors, molecular targeting, or drug-delivery require their functionalization with biologically active and specific molecular ligands. Functionalization protocols often result in a heterogeneous population of particles with a varying density, spatial distribution and orientation of the functional groups on the particle surface. A lack of methods to directly resolve these molecular properties of the particle's surface hampers optimization of functionalization protocols and applications. Here quantitative single-molecule interaction kinetics is used to count the number of ligands on the surface of hundreds of individual nanoparticles simultaneously. By analyzing the waiting-time between single-molecule binding events we quantify the particle functionalization both accurately and precisely for a large range of ligand densities. We observe significant particle-to-particle differences in functionalization which are dominated by the particle-size distribution for high molecular densities, but are substantially broadened for sparsely functionalized particles. From time-dependent studies we find that ligand reorganization on long timescales drastically reduces this heterogeneity, a process that has remained hidden up to now in ensemble-averaged studies. The quantitative single-molecule counting therefore provides a direct route to quantification and optimization of coupling protocols towards molecularly controlled colloidal interfaces.

Strong Plasmon Enhancement of the Saturation Photon Count Rate of Single Molecules.

Abstract: Plasmon resonances have appeared as a promising method to boost the fluorescence intensity of single emitters. However, because research has focused on the enhancement at low excitation intensity, little is known about plasmon-fluorophore coupling near the point where the dye saturates. Here we study plasmon-enhanced fluorescence at a broad range of excitation intensities up to saturation. We adopt a novel DNA-mediated approach wherein dynamic single-molecule binding provides a controlled particle-fluorophore spacing, and dynamic rebinding circumvents artifacts due to photobleaching. We find that near saturation the maximum photon count rate is enhanced by more than 2 orders of magnitude at the optimal particle-fluorophore spacing, even for a dye with a high intrinsic quantum yield. We compare our results to a numerical model taking into account dye saturation. These experiments provide design rules to maximize the photon output of single emitters, which will open the door to studying fast dynamics in real time using single-molecule fluorescence.

Correction to "Strong Plasmon Enhancement of the Saturation Photon Count Rate of Single Molecules".

Abstract: R we published an article in Journal of Physical Chemistry Letters titled “Strong Plasmon Enhancement of the Saturation Photon Count Rate of Single Molecules” (DOI: 10.1021/acs.jpclett.0c00155; publication date: February 19, 2020). Unfortunately, it recently came to our attention that there is a typographical error in the discussion of Figure 6. Following the comments by one of the referees we modified the manuscript regarding the triplet state modifications, but we mistakenly state that the ratio γisc/γT might be at the origin of the mismatch we find in Isat. Obviously, if PCRmax matches the expectation, the ratio γisc/γT also matches the expectation. The text should therefore read that not the ratio γisc/γT but the absolute value of γisc is at the origin of the mismatch. If possible we would also like broaden the argument and include one sentence that an overestimation of the particle−fluorophore spacing might contribute as well because γnr scales more strongly with distance than γr. The paragraph containing the above-mentioned additions and corrections should read as follows: