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This Analysis reports a comparison of current software packages for single-molecule localization in localization-based super-resolution imaging. Performance of the participating software on synthetic, biologically inspired ground-truth data was assessed by multiple criteria. The quality of super-resolution images obtained by single-molecule localization microscopy (SMLM) depends largely on the software used to detect and accurately localize point sources. In this work, we focus on the computational aspects of super-resolution microscopy and present a comprehensive evaluation of localization software packages. Our philosophy is to evaluate each package as a whole, thus maintaining the integrity of the software. We prepared synthetic data that represent three-dimensional structures modeled after biological components, taking excitation parameters, noise sources, point-spread functions and pixelation into account. We then asked developers to run their software on our data; most responded favorably, allowing us to present a broad picture of the methods available. We evaluated their results using quantitative and user-interpretable criteria: detection rate, accuracy, quality of image reconstruction, resolution, software usability and computational resources. These metrics reflect the various tradeoffs of SMLM software packages and help users to choose the software that fits their needs.

Dopamine is a key molecule in neurotransmission and has been known to be responsible for several neurological diseases. Hence, its sensitive and selective detection is important for the early diagnosis of diseases related to abnormal levels of dopamine. In this study, we reported a new cylindrical gold nanoelectrode (CAuNE) platform fabricated via sequential laser interference lithography and electrochemical deposition. Among the fabricated electrodes, CAuNEs with a diameter of 700 nm, 150 s deposited, was found to be the best for electrochemical dopamine detection. According to cyclic voltammetry results, the linear range of the CAuNE-700 nm was 1–100 µM of dopamine with a limit of detection (LOD) of 5.83 µM. Moreover, owing to the homogeneous periodic features of CAuNEs, human neural cells were successfully cultured and maintained for more than 5 days in vitro without the use of any extracellular matrix proteins and dopamine was detectable in the presence of these cells on the electrode. Therefore, we concluded that the developed dopamine sensing platform CAuNE can be used for many applications including early diagnosis of neurological diseases; function tests of dopaminergic neurons derived from various stem cell sources; and toxicity assessments of drugs, chemicals, and nanomaterials on human neuronal cells.

The tumor microenvironment, including stromal cells, surrounding blood vessels and extracellular matrix components, has been defined as a crucial factor that influences the proliferation, drug-resistance, invasion and metastasis of malignant epithelial cells. Among other factors, the communications and interaction between cancer cells and stromal cells have been reported to play pivotal roles in cancer promotion and progression. To investigate these relationships, an on-chip co-culture model was developed to study the cellular interaction between A549-human lung carcinoma cells and MRC-5-human lung epithelial cells in both normal proliferation and treatment conditions. In brief, a co-culture device consisting of 2 individual fluidic chambers in parallel, which were separated by a 100 μm fence was utilized for cell patterning. Microelectrodes arrays were installed within each chamber including electrodes at various distances away from the confrontation line for the electrochemical impedimetric sensing assessment of cell-to-cell influence. After the fence was removed and cell-to-cell contact occurred, by evaluating the impedance signal responses representing cell condition and behavior, both direct and indirect cell-to-cell interactions through conditioned media were investigated. The impact of specific distances that lead to different influences of fibroblast cells on cancer cells in the co-culture environment was also defined.

Super resolution microscopy such as STORM and (F)PALM is now a well known method for biological studies at the nanometer scale. However, conventional imaging schemes based on sparse activation of photo-switchable fluorescent probes have inherently slow temporal resolution which is a serious limitation when investigating live-cell dynamics. Here, we present an algorithm for high-density super-resolution microscopy which combines a sparsity-promoting formulation with a Taylor series approximation of the PSF. Our algorithm is designed to provide unbiased localization on continuous space and high recall rates for high-density imaging and to have orders-of-magnitude shorter run times compared to previous high-density algorithms. We validated our algorithm on both simulated and experimental data and demonstrated live-cell imaging with temporal resolution of 2.5 seconds by recovering fast ER dynamics.

Adoptive cell therapy with chimeric antigen receptor (CAR)-engineered T cells (CAR-Ts) has emerged as an innovative immunotherapy for hematological cancer treatment. However, the limited effect on solid tumors, complex processes, and excessive manufacturing costs remain as limitations of CAR-T therapy. Nanotechnology provides an alternative to the conventional CAR-T therapy. Owing to their unique physicochemical properties, nanoparticles can not only serve as a delivery platform for drugs but also target specific cells. Nanoparticle-based CAR therapy can be applied not only to T cells but also to CAR-natural killer and CAR-macrophage, compensating for some of their limitations. This review focuses on the introduction of nanoparticle-based advanced CAR immune cell therapy and future perspectives on immune cell reprogramming.

C-Reactive protein (CRP) is a biomarker of inflammatory responses and an index for assessing the risk of cardiovascular disease and estimating prognosis. In this study, we constructed a surface-enhanced Raman spectroscopy (SERS) biosensor composed of a multifunctional DNA three-way junction (DNA 3WJ), porous gold nanoplates (pAuNPs), and an Au-Te nanoworm structure for detection of CRP. The pAuNP and Au-Te nanostructures were synthesized by galvanic replacement reactions, and the morphology was confirmed by transmission electron microscopy, scanning electron microscopy, and dynamic light scattering (DLS). To generate the SERS signal, the Au-Te nanostructure was immobilized on an indium-tin oxide substrate, and the thiol-modified CRP aptamer was then self-assembled onto the modified substrate for CRP recognition. To amplify the SERS signal and identify the Raman tag, the multifunctional DNA 3WJ was conjugated with the pAuNPs, and each fragment of 3WJ was functionalized to biotin (pAuNP conjugation), methylene blue (Raman reporter), and CRP aptamer (target binding). The results were confirmed by gel electrophoresis. For conjugation between pAuNPs and DNA 3WJ, avidin was encapsulated in pAuNPs, and the conjugation structure was confirmed by DLS. The fabricated SERS biosensor showed detection limits of 2.23 pM in phosphate-buffered saline and 3.11 pM in diluted human serum. Overall, the proposed biosensor may have potential applications as a SERS biosensor platform.

The unprecedented threat of the highly contagious virus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes exponentially increased infections of coronavirus disease 2019 (COVID-19), highlights the weak spots of the current diagnostic toolbox. In the midst of catastrophe, nanobiosensors offer a new opportunity as an alternative tool to fill a gap among molecular tests, rapid antigen tests, and serological tests. Nanobiosensors surpass the potential of antigen tests because of their enhanced sensitivity, thus enabling us to see antigens as stable and easy-to-access targets. During the first three years of the COVID-19 pandemic, a substantial number of studies have reported nanobiosensors for the detection of SARS-CoV-2 antigens. The number of articles on nanobiosensors and SARS-CoV-2 exceeds the amount of nanobiosensor research on detecting previous infectious diseases, from influenza to SARS-CoV and MERS-CoV. This unprecedented publishing pace also implies the significance of SARS-CoV-2 and the present pandemic. In this review, 158 studies reporting nanobiosensors for detecting SARS-CoV-2 antigens are collected to discuss the current challenges of nanobiosensors using the criteria of point-of-care (POC) diagnostics along with COVID-specific issues. These advances and lessons during the pandemic pave the way for preparing for the post-COVID era and potential upcoming infectious diseases.

Aflatoxin B1 (AFB1) is produced by the Aspergillus flavus and Aspergillus parasiticus group of fungi which is most hepatotoxic and hepatocarcinogenic and occurs as a contaminant in a variety of foods. AFB1 is mutagenic, teratogenic, and causes immunosuppression in animals and is mostly found in peanuts, corn, and food grains. Therefore, novel methodologies of sensitive and expedient strategy are often required to detect mycotoxins at the lowest level. Herein, we report an electrochemical impedance sensor that selectively detects AFB1 at the lowest level by utilizing polyaniline nanofibers (PANI) coated with gold (Au) nanoparticles composite based indium tin oxide (ITO) disk electrodes. The Au-PANI nanocomposites were characterized by scanning electron microscopy (SEM), X-ray diffraction (XRD) spectroscopy, and electrochemical impedance spectroscopy (EIS). The composite electrode exhibited a 14-fold decrement in |Z|1 Hz in comparison with the bare electrode. The Au-PANI acted as an effective sensing platform having high surface area, electrochemical conductivity, and biocompatibility which enabled greater loading deposits of capture antibodies. As a result, the presence of AFB1 was screened with high sensitivity and stability by monitoring the changes in impedance magnitude (|Z|) in the presence of a standard iron probe which was target specific and proportional to logarithmic AFB1 concentrations (CAFB1). The sensor exhibits a linear range 0.1 to 100 ng/mL with a detection limit (3σ) of 0.05 ng/mL and possesses good reproducibility and high selectivity against another fungal mycotoxin, Ochratoxin A (OTA). With regard to the practicability, the proposed sensor was successfully applied to spiked corn samples and proved excellent potential for AFB1 detection and development of point-of-care (POC) disease sensing applications.

Zika virus is a highly infectious virus that is part of the flavivirus group. Precise diagnosis of the Zika virus is significant issue for controlling a global pandemic after the COVID-19 era. For the first time, we describe a zika virus aptamer-based electrical biosensor for detecting Zika virus in human serum. The electrical biosensor composed of a Zika virus aptamer/MXene nanoparticle heterolayer on Au micro-gap electrode (AuMGE)/print circuit board (PCB) system. The Zika virus aptamer was designed to bind the envelope protein of the Zika virus by systematic evolution of ligands by exponential enrichment (SELEX) technique. The binding affinity of the aptamer was determined by fluorescence. For improving the sensor signal sensitivity, Ti3C2Tx MXene was introduced to surface of Au micro-gap electrode (AuMGE). The immobilization process was confirmed by atomic force microscopy (AFM). The prepared aptamer/MXene immobilized on AuMGE can detect the Zika virus through capacitance change according to the target concentration. The capacitance signal from the biosensor increased linearly according to increment of envelope proteins in the human serum. The limit of detection was determined to 38.14 pM, and target proteins could be detected from 100 pM to 10 μM. Thus, the developed electrical aptabiosensor can be a useful tool for Zika virus detection.