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SignificanceThe dual-wedge prism (DWP)-based spectroscopic single-molecule localization microscopy (sSMLM) system offers improved localization precision and adjustable spectral or localization performance, but its nonlinear spectral dispersion presents a challenge. A systematic method can help understand the challenges and thereafter optimize the DWP system’s performance by customizing system parameters to maximize spectral or localization performance for various molecular labels.AimWe developed an MC-based model which predicts the imaging output of the DWP-based sSMLM system given different system parameters.ApproachWe assessed our MC model’s localization and spectral precisions by comparing our simulation against theoretical equations and fluorescent microspheres. Furthermore, we simulated the DWP-based system using beamsplitters of Reflectance (R):Transmittance (T) of R50:T50 and R30:T70 and their tradeoffs.ResultsOur MC simulation showed average deviations of 2.5 nm and 2.1 nm for localization and spectral precisions against theoretical equations; and 2.3 nm and 1.0 nm against fluorescent microspheres. An R30:T70 beamsplitter improved spectral precision by 8% but worsened localization precision by 35% on average compared to an R50:T50 beamsplitter.ConclusionsThe MC model accurately predicted localization precision, spectral precision, spectral peaks, and spectral widths of fluorescent microspheres, as validated by experimental data. Our work enhances the theoretical understanding of DWP-based sSMLM for multiplexed imaging, enabling performance optimization.

By manipulating the spectral dispersion of detected photons, spectroscopic single-molecule localization microscopy (sSMLM) permits concurrent high-throughput single-molecular spectroscopic analysis and imaging. Despite its promising potential, using discrete optical components and managing the delicate balance between spectral dispersion and spatial localization compromise its performance, including nonuniform spectral dispersion, high transmission loss of grating, high optical alignment demands, and reduced precision. We designed a dual-wedge prism (DWP)-based monolithic imaging spectrometer to overcome these challenges. We optimized the DWP for spectrally dispersing focused beam without deviation and with minimal wavefront error. We integrated all components into a compact assembly, minimizing total transmission loss and significantly reducing optical alignment requirements. We show the feasibility of DWP using ray-tracing and numerical simulations. We validated our numerical simulations by experimentally imaging individual nanospheres and confirmed that DWP-sSMLM achieved much improved spatial and spectral precisions of grating-based sSMLM. We also demonstrated DWP-sSMLM in 3D multi-color imaging of cells.

Single-molecule localization microscopy (SMLM) enables the detailed visualization of nuclear pore complexes (NPC) in vitro with sub-20 nm resolution. However, it is challenging to translate the localized coordinates in SMLM images to NPC functions because different algorithms to cluster localizations as individual NPCs can be biased without ground truth for validation. We developed a Monte-Carlo simulation to generate synthetic SMLM images of NPC and used the simulated NPC images as the ground truth to evaluate the performance of six clustering algorithms. We identified HDBSCAN as the optimal clustering algorithm for NPC counting and sizing. Furthermore, we compared the clustering results between the experimental and synthetic data for NUP133, a subunit in the NPC, and found them to be in good agreement.

The roof is the primary heat source for landed buildings since it is exposed to the sun. This will lead to significant heat gain in the attic, causing thermal discomfort for the indoor dwellers and increasing cooling loads. An ideal cool roof system plays an important role in inhibiting excessive heat gain and lowering the cooling load for attic temperature reduction. The experiment was conducted indoors by projecting two 500 W halogen spotlights at each roof model to replace the sunlight. The temperature of roof surface, attic, and moving-air-cavity (MAC) were measured using K-type thermocouples. The variation of temperature versus time for each roof model was compared against the predecessor design and base model. Significantly, with the cool roof model integrating vegetation layer, lightweight foam concrete roof tile, and active MAC with solar-powered fans, the attic temperature remained cool at 26.9 °C with a rate of 0.003 °C/min, 96.77% lower than the based model with reinforced concrete roof. The outstanding performance is due to the inventive cool roof system comprising the ability to minimize the heat gain while circulating the hot air efficiently in keeping the attic cool.

Titanium is one of the biomaterials commonly used for prosthetic devices due to its bio-inert properties. The discovery of titanium dioxide nanotubes (TDNTs) has created a great interest in medical applications such as dental and orthopedic implants. The synthesizing of TDNTs can produce different morphology, sizes and mechanical properties of the nanotubes – depending on the applied method. In this study, an electrochemical anodization method was used for synthesizing the TDNTs. A 100 ml mixture of 99% of ethylene glycol (EG), 1% of deionized water and 1 wt.% of ammonium fluoride (NH4F) was used as the electrolyte of the electrochemical cell. Parameters such as anodization time and the voltage applied were used to alter the morphology of the TDNTs formed. The produced nanotubes were analyzed and characterized using scanning electron microscopy (SEM), X-ray diffraction (XRD) and microhardness tester. The SEM results showed that the formed diameter of nanotubes was mainly affected by the anodizing voltage. The wall thickness was found to be irrelevant to the parameters conducted in this study. The diameter of nanotubes formed with an anodizing voltage of 30, 45 and 60 V have the diameters ranging from 46 nm to 71 nm. All of the TDNTs samples formed have a wall thickness between 11 nm and 13 nm. With the use of EG and NH4F as an electrolyte, the array of TDNTs with honeycomb structure was formed. In general, hardness test showed that the hardness of the nanotubes was inversely proportional with the anodizing time. The anodizing voltage only has little effect on the hardness of the nanotubes. The nanotubes formed by 60 V have about 3 to 5% lower hardness compared to those formed by 30 V for different anodizing times.