Optical Readout Research Articles

Detecting single nanoparticles using fiber-tip nanophotonics

Sensing nano-objects, from nanoparticles to molecules, has become a crucial need in environmental monitoring, medical diagnostics, and drug development. Detection of single particles and molecules is highly desirable, as it provides specific information on size, dynamics, and interactions. Current nanophotonic implementations rely on complex optical readout schemes, limiting their application in the field. Here we demonstrate a nanophotonic fiber-tip sensor with a compact sensor footprint and a simple readout scheme. We leverage advanced design methods to simultaneously achieve a small mode volume V m =0.74(λ/n)3, narrow linewidth Δλ=0.4nm, and a large modulation ΔR≈20% in reflection from the fiber. This unique combination of properties opens the way to sensing weak nanoscale perturbations in the vicinity of the fiber tip. In particular, we experimentally demonstrate the real-time detection of single 50 nm nanoparticles. This opens a route towards real-time sensing of single nanoparticles, and potentially single molecules, in environmental monitoring and diagnostics.

Scaling the Functional Nanopore (FuN) Screen: Systematic Evaluation of Self-Assembling Membrane Peptides and Extension with a K+-Responsive Fluorescent Protein Sensor.

The functional analysis of protein nanopores is typically conducted in planar lipid bilayers or liposomes exploiting high-resolution but low-throughput electrical and optical read-outs. Yet, the reconstitution of protein nanopores in vitro still constitutes an empiric and low-throughput process. Addressing these limitations, nanopores can now be analyzed using the functional nanopore (FuN) screen exploiting genetically encoded fluorescent protein sensors that resolve distinct nanopore-dependent Ca2+ in- and efflux patterns across the inner membrane of Escherichia coli. With a primary proof-of-concept established for the S2168 holin, and thereof based recombinant nanopore assemblies, the question arises to what extent alternative nanopores can be analyzed with the FuN screen and to what extent alternative fluorescent protein sensors can be adapted. Focusing on self-assembling membrane peptides, three sets of 13 different nanopores are assessed for their capacity to form nanopores in the context of the FuN screen. Nanopores tested comprise both natural and computationally designed nanopores. Further, the FuN screen is extended to K+-specific fluorescent protein sensors and now provides a capacity to assess the specificity of a nanopore or ion channel. Finally, a comparison to high-resolution biophysical and electrophysiological studies in planar lipid bilayers provides an experimental benchmark for future studies.

A pH-Responsive Topological Switch Based on a DNA Quadruplex-Duplex Hybrid.

A guanine-rich oligonucleotide based on a human telomeric sequence but with the first three-nucleotide intervening stretch replaced by a putative 15-nucleotide hairpin-forming sequence shows a pH-dependent folding into different quadruplex-duplex hybrids in a potassium containing buffer. At slightly acidic pH, the quadruplex domain adopts a chair-type conformation. Upon increasing the pH, a transition with a midpoint close to neutral pH to a major and minor (3+1) hybrid topology with either a coaxially stacked or orthogonally oriented duplex stem-loop occurs. NMR-derived high-resolution structures reveal that an adenine protonation is prerequisite for the formation of a non-canonical base quartet, capping the outer G-tetrad at the quadruplex-duplex interface and stabilizing the antiparallel chair conformation in an acidic environment. Being directly associated with interactions at the quadruplex-duplex interface, this unique pH-dependent topological transition is fully reversible. Coupled with a conformation-sensitive optical readout demonstrated as a proof of concept using the fluorescent dye thiazole orange, the present quadruplex-duplex hybrid architecture represents a potentially valuable pH-sensing system responsive in a physiological pH range of 7±1.

Elucidating Singlet-Fission-Born Multiexciton Dynamics via Molecular Engineering: A Dilution Principle Extended to Quintet Triplet Pair.

Multiexciton in singlet exciton fission represents a critical quantum state with significant implications for both solar cell applications and quantum information science. Two distinct fields of interest explore contrasting phenomena associated with the geminate triplet pair: one focusing on the persistence of long-lived correlation and the other emphasizing efficient decorrelation. Despite the pivotal nature of multiexciton processes, a comprehensive understanding of their dependence on the structural and spin properties of materials is currently lacking in experimental realizations. To address this gap in knowledge, molecular engineering was employed to modify the TIPS-tetracene structures, enabling an investigation of the structure-property relationships in spin-related multiexciton processes. In lieu of the time-resolved electron paramagnetic resonance technique, two time-resolved magneto-optical spectroscopies were implemented for quantitative analysis of spin-dependent multiexciton dynamics. The utilization of absorption and fluorescence signals as complementary optical readouts, in the presence of a magnetic field, provided crucial insights into geminate triplet pair dynamics. These insights encompassed the duration of multiexciton correlation and the involvement of the spin state in multiexciton decorrelation. Furthermore, simulations based on our kinetic models suggested a role for quintet dilution in multiexciton dynamics, surpassing the singlet dilution principle established by the Merrifield model. The integration of intricate model structures and time-resolved magneto-optical spectroscopies served to explicitly elucidate the interplay between structural and spin properties in multiexciton processes. This comprehensive approach not only contributes to the fundamental understanding of these processes but also aligns with and reinforces previous experimental studies of solid states and theoretical assessments.

Wireless Light-Emitting Electrode Arrays for the Evaluation of Electrocatalytic Activity.

Water splitting has become a sustainable and clean alternative for hydrogen production. Commonly, the efficiency of such reactions is intimately related to the physico-chemical properties of the catalysts that constitute the electrolyzer. Thus, the development of simple and fast methods to evaluate the electrocatalytic efficiency of an electrolyzer is highly required. In this work, we present an unconventional method based on the combination of bipolar electrochemistry and light-emitting diodes, which allows the evaluation of the electrocatalytic performance of the two types of catalysts, composing an electrolyzer, namely for oxygen and hydrogen evolution reactions, respectively. The integrated light emission of the diode acts as an optical readout of the electrocatalytic information, which simultaneously depends on the composition of the anode and the cathode. The electrocatalytic activity of Au, Pt, and Ni electrodes, connected to the LED in multiple anode/cathode configurations, towards the water splitting reactions has been evaluated. The efficiency of the electrolyzer can be represented in terms of the onset electric field (ϵonset) for light emission, obtaining variations that are in agreement with data reported with conventional electrochemistry. This work introduces a straightforward method for evaluating electrocatalysts and underscores the importance of material characterization in developing efficient electrolyzers for hydrogen production.

POLARISED views and FRETting about probe modulation assays: Learning from High Throughput Screening

Fluorescent probe modulation assays are a widely used approach to monitor displacement or stabilisation of fluorescently labelled tool ligands by test compounds. These assays allow an optical read-out of probe-receptor binding and can be used to detect compounds that compete with the labelled ligand, either directly or indirectly. Probes for both orthosteric and allosteric sites are often employed. The method can also be used to identify test compounds that may stabilise the ternary complex, offering an opportunity to discover novel molecular glues. The utility of these fluorescence-based assays within high-throughput screening has been facilitated by the use of streptavidin labelled terbium as a donor and access to a range of different acceptor fluorophores. During 2023, the High-throughput Screening group at AstraZeneca carried out 8 high-throughput screens using these approaches. In this manuscript we will present the types of assays used, an overview of the timelines for assay development and screening, the application of orthogonal artefact methods to aid hit finding and the results of the screens in terms of hit rate and the number of compounds identified with IC50 values of better than 30 µM. Learning across the development, execution and analysis of these screens will be presented.

Abstract 5371: Multimodal CRISPR activation screens reveal new mechanisms to sensitize cancer cells to targeted T cell mediated elimination

Abstract Cell autonomous resistance mechanisms can protect cancer cells from targeted T cell elimination, resulting in resistance to a broad range of immunotherapies. High-throughput gene knockout and gene inhibition screens have been instrumental in identifying key drivers of such mechanisms, yet often uncovering gene activity that is essential but not sufficient for response. Here we set out to leverage gene activation as an orthogonal approach to identify novel regulators of the cancer-immune interface. First, we performed CRISPR activation screens in melanoma cells co-cultured with antigen-specific primary CD8+ T cells. Our findings revealed a collection of novel genes that substantially sensitize melanoma cells to targeted T cell-driven elimination when activated, including DNA damage and unfolded protein response genes (e.g., ETS1, TSPYL2), Wnt ligands (e.g., WNT3A, WNT1), and others that are deleterious only in combination with targeting T cells. Among the resistance genes we found regulators of insulin signaling (e.g., HGF, TCF7L2) and numerous glycoproteins (e.g., PDPN, CD44). Second, we validated top novel hits in 2D and in 3D spheroid models, using optical readouts to investigate their impact on cancer cell elimination and cancer-T-cell spatial dynamics. Third, we performed a Perturb-seq screen to stratify dozens of hits into functional groups based on the transcriptional readouts in the cancer cells. Lastly, we developed an in situ Perturb-Seq method, where we combine high-plex spatial transcriptomic readouts at subcellular resolution with in situ detection of genetically modified cells, and applied it to track the impact of the (de)sensitizing perturbations not only on the cancer cells themselves, but also on T cell recruitment and suppression. Taken together, our study provides a catalog of immune evasion mechanisms and an array of novel targets, demonstrating that synthetic gene activation can diminish immune evasion and open novel avenues for development of agonist-based and RNA-based interventions. Citation Format: Reece V. Akana, Chang Sun, YoungMin Kim, Yuxin Cai, Subin Kim, Jeehyun Yoe, Olivia Laveroni, Livnat Jerby-Arnon. Multimodal CRISPR activation screens reveal new mechanisms to sensitize cancer cells to targeted T cell mediated elimination [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5371.

Nanophotonic resistive switch based on tapered copper-silicon structure with low power and high extinction ratio

A non-volatile nanophotonic resistive switch in tapered copper-on-silicon structure is proposed with a potential of electrical writing and optical readout. Incorporating tapered hybrid plasmonic structure in the device enables variable areas of the density of conduction filaments in the direction of optical propagation causing enhanced interaction of light with the lossy metal filaments. The tapered Cu-SiO2-Si is shown to have large optical extinction ratio where high mobility of copper ions in thin SiO2 leads to the low power operation. Optical readout of the resistive switching is demonstrated in the 10 nm thin ion conducting layer (SiO2) at a 1550 nm wavelength with well-defined hysteresis curve. Switching between off and on states is attained by voltage induced annihilation/formation of the nanoscale metal filament inside a thin layer of SiO2 sandwiched between copper and silicon. An optical extinction ratio of 18 dB is reported for a 30 μm long device. In addition, the proposed nanophotonic switch exhibits an intrinsic stochastic property of set voltage reduction in each set-reset cycle. High extinction ratio, reduced set voltage, low power consumption, and non-volatility make the device excellent choice for applications in realizing ultra-compact on-chip devices for optical switching, modulation, and neuromorphic computing.

Low-power, agile electro-optic frequency comb spectrometer for integrated sensors.

Sensing platforms based upon photonic integrated circuits have shown considerable promise; however, they require corresponding advancements in integrated optical readout technologies. Here, we present an on-chip spectrometer that leverages an integrated thin-film lithium niobate modulator to produce a frequency-agile electro-optic frequency comb for interrogating chip-scale temperature and acceleration sensors. The chirped comb process allows for ultralow radiofrequency drive voltages, which are as much as seven orders of magnitude less than the lowest found in the literature and are generated using a chip-scale, microcontroller-driven direct digital synthesizer. The on-chip comb spectrometer is able to simultaneously interrogate both an on-chip temperature sensor and an off-chip, microfabricated optomechanical accelerometer with cutting-edge sensitivities of ≈5µK⋅Hz−1/2 and ≈130µm⋅s−2⋅Hz−1/2, respectively. This platform is compatible with a broad range of existing photonic integrated circuit technologies, where its combination of frequency agility and ultralow radiofrequency power requirements are expected to have applications in fields such as quantum science and optical computing.

Hexagonal Boron Nitride Spacers for Fluorescence Imaging of Biomolecules

AbstractFluorescence imaging is an invaluable tool to investigate biomolecular dynamics, mechanics, and interactions in aqueous environments. Two‐dimensional materials offer large‐area, atomically smooth surfaces for wide‐field biomolecule imaging. Despite the success of graphene for on‐chip biosensing and biomolecule manipulation, its strong fluorescence‐quenching properties pose a challenge for biomolecular investigations that are based on direct optical readouts. Here, we employ few‐layer hexagonal boron nitride (hBN) as a precisely tailorable fluorescence spacer between labelled lipid membranes and graphene substrates. By stacking high‐quality hBN crystals in the 10–20 nm thickness range on monolayer graphene, we observe distance‐dependent fluorescence intensity variations. Remarkably, with hBN spacers as thin as 20 nm, the fluorescence intensity is comparable to bare SiO2/Si substrates, while the intensity was reduced to 60 % and 80 % with ~10 nm and ~16 nm hBN thicknesses respectively. We confirm that pre‐determined hBN thicknesses can be employed to control the non‐radiative energy transfer properties of graphene, with fluorescence quenching following a d−4 distance‐dependent behaviour. This seamless integration of electronically active and dielectric van der Waals materials into vertical heterostructures enables multifunctional platforms addressing the manipulation, localization, and visualization of biomolecules for fundamental biophysics and biosensing applications.

Compact silicon photonic Mach-Zehnder modulators for high-energy physics

The characterization of compact non-traveling-wave Mach-Zehnder modulators for optical readout in high-energy physics experiments is reported to provide power-efficient alternatives to conventional traveling-wave devices and a more resilient operation compared to ring modulators. Electro-optical small-signal and large-signal measurements showcase the performances of custom NTW-MZMs designed and fabricated in iSiPP50G IMEC's technology in the framework of INFN's FALAPHEL project. Bit-error-rate results demonstrate their potential suitability for optical links up to 25 Gb/s when equipped with either conventional deep-etched or radiation-hardened shallow-etched free-carrier-based phase shifters.

Optical measurement of glutamate release robustly reports short-term plasticity at a fast central synapse.

Recently developed fluorescent neurotransmitter indicators have enabled direct measurements of neurotransmitter in the synaptic cleft. Precise optical measurements of neurotransmitter release may be used to make inferences about presynaptic function independent of electrophysiological measurements. Here, we express iGluSnFR, a genetically encoded glutamate reporter in mouse spiral ganglion neurons to compare electrophysiological and optical readouts of presynaptic function and short-term synaptic plasticity at the endbulb of Held synapse. We show iGluSnFR robustly and approximately linearly reports glutamate release from the endbulb of Held during synaptic transmission and allows assessment of short-term plasticity during high-frequency train stimuli. Furthermore, we show that iGluSnFR expression slightly alters the time course of spontaneous postsynaptic currents, but is unlikely to impact measurements of evoked synchronous release of many synaptic vesicles. We conclude that monitoring glutamate with optical sensors at fast and large central synapses like the endbulb of Held is feasible and allows robust quantification of some, but not all aspects of glutamate release.

Infrared vertical external cavity surface emitting laser threshold magnetometer

Nitrogen-vacancy (NV) centers have considerable promise as high sensitivity magnetometers; however, they are commonly limited by inefficient collection and low contrasts. Laser threshold magnetometry (LTM) enables efficient collection and high contrasts, providing a path toward higher sensitivity magnetometry. We demonstrate an infrared LTM using an ensemble of NV centers in a single-crystal diamond plate integrated into a vertical external cavity surface emitting laser. The laser was tuned to the spin-dependent absorption line of the NV centers, allowing for optical readout by monitoring the laser output power. We demonstrate a magnetic sensitivity of 7.5 nT/Hz in the frequency range between 10 and 50 Hz. Furthermore, the contrast and the projected photon shot noise limited (PSNL) sensitivity are shown to improve significantly by operating close to the lasing threshold, achieving 18.4% and 26.6 pT/Hz near the threshold. In addition, an unexpected saturable absorption phenomenon was observed near the threshold, which enhanced the contrast and projected PSNL sensitivity.

Recent advances in bioluminescent probes for neurobiology.

Bioluminescence is a popular modality for imaging in living organisms. The platform relies on enzymatically (luciferase) generated light via the oxidation of small molecule luciferins. Since no external light is needed for photon production, there are no concerns with background autofluorescence or photobleaching over time-features that have historically limited other optical readouts. Bioluminescence is thus routinely used for longitudinal tracking across whole animals. Applications in the brain, though, have been more challenging due to a lack of sufficiently bioavailable, bright, and easily multiplexed probes. Recent years have seen the development of designer luciferase and luciferin pairs that address these issues, providing more sensitive and real-time readouts of biochemical features relevant to neurobiology. This review highlights many of the advances in bioluminescent probe design, with a focus on the small molecule light emitter, the luciferin. Specific efforts to improve luciferin pharmacokinetics and tissue-penetrant emission are covered, in addition to applications that such probes have enabled. The continued development of improved bioluminescent probes will aid in illuminating critical neurochemical processes in the brain.

Wide-field Fourier magnetic imaging with electron spins in diamond

Wide-field magnetic imaging based on nitrogen-vacancy (NV) centers in diamond has been shown the applicability in material and biological science. However, the spatial resolution is limited by the optical diffraction limit (>200 nm) due to the optical real-space localization and readout of NV centers. Here, we report the wide-field Fourier magnetic imaging technique to improve spatial resolution beyond the optical diffraction limit while maintaining the large field of view. Our method relies on wide-field pulsed magnetic field gradient encoding of NV spins and Fourier transform under pixel-dependent spatial filters. We have improved spatial resolution by a factor of 20 compared to the optical resolution and demonstrated the wide-field super-resolution magnetic imaging of a gradient magnetic field. This technique paves a way for efficient magnetic imaging of large-scale fine structures at the nanoscale.

"Four-In-One" Multifunctional Dandelion-Like Gold@platinum Nanoparticles-Driven Multimodal Lateral Flow Immunoassay.

The traditional lateral flow immunoassay (LFIA) with a single signal output mode may encounter challenges such as low sensitivity, poor detection range, and susceptibility to external interferences. These limitations hinder its ability to meet the growing demand for advanced LFIA. To address these issues, the rational development of multifunctional labels for multimodal LFIA emerges as a promising strategy. Herein, this study reports a multimodal LFIA using "four-in-one" multifunctional dandelion-like gold@platinum nanoparticles (MDGP). The inherent properties of MDGP, such as the broad absorption spectrum, porous dandelion-like nanostructure, and bimetallic composition with gold and platinum, endow them with capacities in dual spectral-overlapped fluorescence quenching, optical readout, catalytic activity, and photothermal effect. Benefiting from their multifunctional properties, the MDGP-LFIA enables multimodal outputs including fluorescent, colorimetric, and photothermal signals. This multimodal MDGP-LFIA allows for the detection of acetamiprid at a range of 0.01-50ngmL-1, with the lowest qualitative and quantitative detection results of 0.5 and 0.008ngmL-1, respectively, significantly better than the traditional gold nanoparticles-based LFIA. The diversity, complementarity, and synergistic effect of integrated output signals in this multimodal MDGP-LFIA improve the flexibility, practicability, and accuracy of detection, holding great promise as a point-of-care testing platform in versatile application scenarios.

Sub-Diffraction Readout Method of High-Capacity Optical Data Storage Based on Polarization Modulation.

The big data era demands an efficient and permanent data storage technology with the capacity of PB to EB scale. Optical data storage (ODS) offers a good candidate for long-lifetime storage, as the developing far-field super-resolution nanoscale writing technology improves its capacity to the PB scale. However, methods to efficiently read out this intensive ODS data are still lacking. In this paper, we demonstrate a sub-diffraction readout method based on polarization modulation, which experimentally achieves the sub-diffraction readout on Disperse Red 13 thin film with a resolution of 500 nm, exceeding the diffraction limit by 1.2 times (NA = 0.5). Differing from conventional binary encoding, we propose a specific polarization encoding method that enhances the capacity of ODS by 1.5 times. In the simulation, our method provides an optical data storage readout resolution of 150 nm, potentially to 70 nm, equivalent to 1.1 PB in a DVD-sized disk. This sub-diffraction readout method has great potential as a powerful readout tool for next-generation optical data storage.

Real-time tracking of coherent oscillations of electrons in a nanodevice by photo-assisted tunnelling

Coherent collective oscillations of electrons excited in metallic nanostructures (localized surface plasmons) can confine incident light to atomic scales and enable strong light-matter interactions, which depend nonlinearly on the local field. Direct sampling of such collective electron oscillations in real-time is crucial to performing petahertz scale optical modulation, control, and readout in a quantum nanodevice. Here, we demonstrate real-time tracking of collective electron oscillations in an Au bowtie nanoantenna, by recording photo-assisted tunnelling currents generated by such oscillations in this quantum nanodevice. The collective electron oscillations show a noninstantaneous response to the driving laser fields with a T2 decay time of nearly 8 femtoseconds. The contributions of linear and nonlinear electron oscillations in the generated tunnelling currents were precisely determined. A phase control of electron oscillations in the nanodevice is illustrated. Functioning in ambient conditions, the excitation, phase control, and read-out of coherent electron oscillations pave the way toward on-chip light-wave electronics in quantum nanodevices.

Convergence of Surface-Enhanced Raman Scattering with Molecular Diagnostics: A Perspective on Future Directions.

Molecular diagnostics (MD) is widely employed in multiple scientific disciplines, such as oncology, pathogen detection, forensic investigations, and the pharmaceutical industry. Techniques such as polymerase chain reaction (PCR) revolutionized the rapid and accurate identification of nucleic acids (DNA, RNA). More recently, CRISPR and its CRISPR-associated protein (Cas) have been a ground-breaking discovery that is the latest revolution in molecular biology, including MD. Surface-enhanced Raman scattering (SERS) is a very attractive alternative to fluorescence as the currently most widely used optical readout in MD. In this Perspective, milestones in the development of MD, SERS-PCR, and next-generation approaches to MD, such as Specific High-Sensitivity Enzymatic Reporter UnLOCKing (SHERLOCK) and DNA Endonuclease-Targeted CRISPR Trans Reporter (DETECTR), are briefly summarized. Our perspective on the future convergence of SERS with MD is focused on SERS-based CRISPR/Cas (SERS-CRISPR) since we anticipate many promising applications in this rapidly emerging field. We predict that major future developments will exploit the advantages of real-time monitoring with the superior brightness, photostability, and spectral multiplexing potential of SERS nanotags in an automated workflow for rapid assays under isothermal, amplification-free conditions.

Facile and Ultrasensitive Food Allergen Quantification Using Microzone Paper-Based Mass Spectrometric Immunoassay.

Highly sensitive and rapid measurement of food allergens is essential to avoid unanticipated food allergies and to determine whether cross-contamination occurs in the food industry. Commercial immunoassay kits offer high specificity and convenience for allergen detection but still suffer limited quantitative sensitivity, accuracy, and stability based on the optical readout. In this work, a paper-based mass spectrometric immunoassay platform was constructed to achieve facile and highly sensitive quantification of peanut allergen, which combined the advantages of good specificity and accurate quantification from mass spectrometry and simplicity from a paper-based immunoassay. In this platform, a novel quaternary ammonium-based mass tag and a paper chip with a microzone were designed and developed, contributing to a large signal enhancement. This method was able to detect Ara h1 with a linear range of 0.1-100 ng mL-1 and a detection limit of 0.08 ng mL-1 in milk matrices. It has also been successfully applied to the accurate quantification of Ara h1 in six milk-related beverages, two biscuits, and two candy bars with complicated matrices and presented a low-concentration quantitation capability. This method gives a new type of mass spectrometric immunoassay for rapid and ultrasensitive allergen regulation in the food industry and for individual allergen differentiation research.