Analysis of Fluid Replacement in Two Fluidic Chambers for Oblique-Incidence Reflectivity Difference (OI-RD) Biosensor.

Optical biosensors have a significant impact on various aspects of our lives. In many applications of optical biosensors, fluidic chambers play a crucial role in facilitating controlled fluid delivery. It is essential to achieve complete liquid replacement in order to obtain accurate and reliable results. However, the configurations of fluidic chambers vary across different optical biosensors, resulting in diverse fluidic volumes and flow rates, and there are no standardized guidelines for liquid replacement. In this paper, we utilize COMSOL Multiphysics, a finite element analysis software, to investigate the optimal fluid volume required for two types of fluidic chambers in the context of the oblique-incidence reflectivity difference (OI-RD) biosensor. We found that the depth of the fluidic chamber is the most crucial factor influencing the required liquid volume, with the volume being a quadratic function of the depth. Additionally, the required fluid volume is also influenced by the positions on the substrate surface bearing samples, while the flow rate has no impact on the fluid volume.

[Bioinformatics database and production layout visual analysis platform of medicinal plants:a case study of Dao-di medicinal materials in Sichuan province].

In the new stage of trans-omics and trans-subjects for medicinal plants, it is an urgent need to integrate big data, provide interactive applications, and form a unified and multi-level research system and big data platform. Dao-di medicinal material, as an important source of medicinal plants, is a unique quality concept and comprehensive standard of tranditional Chinese medicine(TCM). Several databases have been developed in China and abroad, such as the Encyclopedia of Traditional Chinese Medicine(ETCM) and the Global Pharmacopoeia Genome Database(GPGD). Yet, most databases do not provide multi-dimensional data, including geographic data, phenotype data, compound data, and genetic data. Sichuan, known as the hometown of TCM therapies and the treasure trove of TCM, is the most representative region of medicinal plant diversity in China. According to the latest data of the fourth national survey of TCM resources, there are more than 8 000 TCM and 86 Dao-di medicinal materials in Sichuan province. Based on resource census data and relevant achievements, this study constructed the bioinformatics database of medicinal plants and the visual analysis platform of production layout by taking the Dao-di medicinal materials in Sichuan province as an example, covering geographic data, phenotype data, compound data, and genetic data. It effectively integrates multi-dimensional data of Dao-di medicinal materials and provides different levels of data interaction applications. The platform is the first large-scale multi-dimensional database and visual platform of Dao-di medicinal materials in Sichuan province, which serves as an essential resource for germplasm resources identification, decomposition of biosynthetic pathways, molecular breeding of varieties and provides medicinal plant resource information and data support for development and utilization of medicinal plants in China and abroad.

Early Detection of Cervical Cancer by Fluorescence Lifetime Imaging Microscopy Combined with Unsupervised Machine Learning.

Cervical cancer has high morbidity and mortality rates, affecting hundreds of thousands of women worldwide and requiring more accurate screening for early intervention and follow-up treatment. Cytology is the current dominant clinical screening approach, and though it has been used for decades, it has unsatisfactory sensitivity and specificity. In this work, fluorescence lifetime imaging microscopy (FLIM) was used for the imaging of exfoliated cervical cells in which an endogenous coenzyme involved in metabolism, namely, reduced nicotinamide adenine dinucleotide (phosphate) [NAD(P)H], was detected to evaluate the metabolic status of cells. FLIM images from 71 participants were analyzed by the unsupervised machine learning method to build a prediction model for cervical cancer risk. The FLIM method combined with unsupervised machine learning (FLIM-ML) had a sensitivity and specificity of 90.9% and 100%, respectively, significantly higher than those of the cytology approach. One cancer recurrence case was predicted as high-risk several months earlier using this method as compared to using current clinical methods, implying that FLIM-ML may be very helpful for follow-up cancer care. This study illustrates the clinical applicability of FLIM-ML as a detection method for cervical cancer screening and a convenient tool for follow-up cancer care.

Application of Dual-Enhanced Surface-Enhanced Raman Scattering Probe Technology in the Diagnosis of Tumor Cells in Vitro.

With the development of precision medicine, antigen/antibody-targeted therapy has brought great hope to tumor patients; however, the migration of tumor cells, especially a small number of cells flowing into blood or other tissues, remains a clinical challenge. In particular, it is difficult to use functional gold nanomaterials for targeted clinical tumor diagnosis while simultaneously obtaining stable and highly sensitive Raman signals. Therefore, we developed a detection method for functional Au Nanostars (AuNSs) with dual signal enhancement that can specifically track location and obtain high-intensity surface-enhanced Raman scattering (SERS) signals. First, AuNSs with specific optical properties were synthesized and functionalized. The Raman dye 4-mercapto-hydroxybenzoic acid and polyethylene glycol were coupled with the tumor marker, epidermal growth factor receptor, to obtain the targeted SERS probes. In addition, a detection chip was prepared for Raman detection with physical enhancement, exhibiting a 40-times higher signal intensity than that of quartz glass. This study combines physical enhancement and SERS enhancement technologies to achieve dual enhancement, enabling the detection of a highly sensitive and stable Raman signal; this has potential clinical value for antigen/antibody-targeted tumor diagnosis and treatment.

Visible Particle Identification Using Raman Spectroscopy and Machine Learning.

Visible particle identification is a crucial prerequisite step for process improvement and control during the manufacturing of injectable biotherapeutic drug products. Raman spectroscopy is a technology with several advantages for particle identification including high chemical sensitivity, minimal sample manipulation, and applicability to aqueous solutions. However, considerable effort and experience are required to extract and interpret Raman spectral data. In this study, we applied machine learning algorithms to analyze Raman spectral data for visible particle identification in order to minimize expert support and improve data analysis accuracy. We manually prepared ten types of particle standard solutions to simulate the particle types typically observed during manufacturing and established a Raman spectral library with accurate peak assignments for the visible particles. Five classification algorithms were trained using visible particle Raman spectral data. All models had high prediction accuracy of >98% for all types of visible particles. Our results demonstrate that the combination of Raman spectroscopy and machine learning can provide a simple and accurate data analysis approach for visible particle identification.

Quantifying tagged mRNA export flux via nuclear pore complexes in single live cells.

The mRNA export flux through nuclear pore complexes (NPC) changes under DNA manipulation and hence affects protein translation. However, monitoring the flux of a specific mRNA in single live cell is beyond reach of traditional techniques. We developed a fluorescence-based detection method for measuring the export flux of mRNA through NPC in single live cell using a snapshot image, which had been tested on exogenous genes' expression in HeLa cells, with transfection or infection, and endogenous genes' expression in yeast cells, during incubation and carbon catabolite repression. With its speediness, explicitness and noninvasiveness, we believe that it would be valuable in direct monitoring of gene behavior, and the understanding of gene regulation at a single cell level.

miR-636 inhibits EMT, cell proliferation and cell cycle of ovarian cancer by directly targeting transcription factor Gli2 involved in Hedgehog pathway.

BACKGROUND: Hedgehog (Hh) signaling pathway, which is essential for cell proliferation and differentiation, is noted to be aberrantly activated in tumor from increasing studies in recent years. MicroRNAs (miRNAs) as an important non-coding RNA in cells have been proven to possess a regulatory role specific to the Hh signaling pathway. Here, in vitro and in vivo cellular/molecular experiments were adopted to clarify the regulatory mechanism linking miR-636 to the Hh signaling pathway in ovarian cancer (OVC). METHODS: Protein-protein interaction analysis was performed to identify the hub gene in the Hh pathway. TargetScan database was used to predict the potential upstream regulators for Gli2. qRT-PCR was performed to test the expression of miR-636, while Western blot was conducted to detect the expression of proteins related to the Hh pathway and epithelial-mesenchymal transition (EMT). For cell functional experiments, HO-8910PM OVC cell line was used. MTT assay and wound healing assay were used to measure the effect of miR-636 on cell proliferation and migration. Flow cytometry was carried out to examine the effect of miR-636 on cell cycle, and Western blot was used to identify the change in expression of Hh and EMT-related proteins. Dual-luciferase reporter gene assay was implemented to detect the targeting relationship between miR-636 and Gli2. Xenotransplantation models were established for in vivo examination. RESULTS: Gli2 was identified as the hub gene of the Hh pathway and it was validated to be regulated by miR-636 based on the data from TargetScan and GEO databases. In vitro experiments discovered that miR-636 was significantly lowly expressed in OVC cell lines, and overexpressing miR-636 significantly inhibited HO-8910PM cell proliferation, migration and induced cell cycle arrest in G0/G1 phase, while the inhibition of miR-636 caused opposite results. Dual-luciferase reporter gene assay revealed that Gli2 was the target gene of miR-636 in OVC. Besides, overexpressed miR-636 decreased protein expression of Gli2, and affected the expression of proteins related to the Hh signaling pathway and EMT. Rescue experiments verified that overexpression of Gli2 reversed the inhibitory effect of miR-636 on HO-8910PM cell proliferation and migration, and attenuated the blocking effect of miR-636 on cell cycle. The xenotransplantation experiment suggested that miR-636 inhibited cell growth of OVC by decreasing Gli2 expression. Besides, overexpressing Gli2 potentiated the EMT process of OVC cells via decreasing E-cadherin protein expression and increasing Vimentin protein expression, and it reversed the inhibitory effect of miR-636 on OVC cell proliferation in vivo. CONCLUSION: miR-636 mediates the activation of the Hh pathway via binding to Gli2, thus inhibiting EMT, suppressing cell proliferation and migration of OVC. TRIAL REGISTRATION: The experimental protocol was established, according to the ethical guidelines of the Helsinki Declaration and was approved by the Human Ethics Committee of The Second Affiliated hospital of Zhejiang University School of Medicine (IR2019001235). Written informed consent was obtained from individual or guardian participants.

Dearomatization of 2,3-Disubstituted Indoles via 1,8-Addition of Propargylic (Aza)-para-Quinone Methides.

Dearomatization of indole is a useful strategy to access indolimines: a motif widely exists in biologically active molecules and natural products. Herein, an efficient method for the dearomatization of 2,3-disubstituted indoles to generate diverse indolimines with tetrasubstituted allenes is described. This work accomplishes dearomatization of 2,3-disubstituted indoles through 1,8-addition of (aza)-para-quinone methides, which are generated in situ from propargylic alcohols. A series of synthetically useful indolimines containing quaternary carbon centers and tetrasubstituted allenes can be accessed in good yields (up to 99%). Additionally, the separability of product isomers, diversified product transformations, and easy scale-up of the reaction demonstrate the potential application of this method.

Post-transcriptional adaptation of the aquatic plant Spirodela polyrhiza under stress and hormonal stimuli.

The Lemnaceae family comprises aquatic plants of angiosperms gaining attention due to their utility in wastewater treatment, and rapid production of biomass that can be used as feed, fuel, or food. Moreover, it can serve as a model species for neotenous growth and environmental adaptation. The latter properties are subject to post-transcriptional regulation of gene expression, meriting investigation of how miRNAs in Spirodela polyrhiza, the most basal and most thoroughly sequenced member of the family, are expressed under different growth conditions. To further scientific understanding of its capacity to adapt to environmental cues, we measured miRNA expression and processing of their target sequences under different temperatures, and in the presence of abscisic acid, copper, kinetin, nitrate, and sucrose. Using two small RNA sequencing experiments and one degradome sequencing experiment, we provide evidence for 108 miRNAs. Sequencing cleaved mRNAs validated 42 conserved miRNAs with 83 targets and 24 novel miRNAs regulating 66 targets and created a list of 575 predicted and verified targets. These analyses revealed condition-induced changes in miRNA expression and cleavage activity, and resulted in the addition of stringently reviewed miRNAs to miRBase. This combination of small RNA and degradome sequencing provided not only high confidence predictions of conserved and novel miRNAs and targets, but also a view of the post-transcriptional regulation of adaptations. A unique aspect is the role of miR156 and miR172 expression and activity in its clonal propagation and neoteny. Additionally, low levels of 24 nt sRNAs were observed, despite the lack of recent retrotransposition.

Super-resolution mapping of scaffold nucleoporins in the nuclear pore complex.

The nuclear pore complex (NPC), composed of ∼30 different nucleoporins (Nups), is one of the largest supramolecular structures in eukaryotic cells. Its octagonal ring scaffold perforates the nuclear envelope and features a unique molecular machinery that regulates nucleocytoplasmic transport. However, the precise copy number and the spatial location of each Nup in the native NPC remain obscure due to the inherent difficulty of counting and localizing proteins inside of the sub-micrometer supramolecular complex. Here, we combined super-resolution single-point edge-excitation subdiffraction (SPEED) microscopy and nanobody-specific labeling to reveal the spatial distribution of scaffold Nups within three separate layers in the native NPC with a precision of ∼3 nm. Our data reveal both the radial and axial spatial distributions for Pom121, Nup37 and Nup35 and provide evidence for their copy numbers of 8, 32 and 16, respectively, per NPC. This approach can help pave the path for mapping the entirety of Nups in native NPCs and also other structural components of macromolecular complexes.

Transcriptomic Profiling and Gene Disruption Revealed that Two Genes Related to PUFAs/DHA Biosynthesis May be Essential for Cell Growth of Aurantiochytrium sp.

Aurantiochytrium sp. PKU#SW7 is a thraustochytrid strain that was found to exhibit high potential for docosahexaenoic acid (DHA, C22:6n-3) production. In this work, the transcriptome of Aurantiochytrium sp. PKU#SW7 was analyzed for the study of genes involved in basic metabolic functions and especially in the mechanisms of DHA biosynthesis. Sequence annotation and functional analysis revealed that the strain contains components of fatty acid synthesis (FAS) and polyketide synthase (PKS) pathways. Fatty acid desaturases and elongases were identified as components of FAS pathway, whilst key components of PKS pathway were also found in the cDNA library. The relative contribution of the two pathways to the synthesis of DHA was unknown, as both pathways appeared to be lacking full complement of genes for standalone synthesis of DHA. Further analysis of two putative genes encoding the very-long-chain (3R)-3-hydroxyacyl-CoA dehydratase and dehydrase/isomerase involved in FAS and PKS pathways, respectively, revealed that under various salinity conditions, their relative expression levels changed corresponding to the variation of DHA content in Aurantiochytrium sp. Independent knock outs of these genes in Aurantiochytrium sp. resulted in poor cell growth, probably due to little or no intracellular DHA accumulation. Hence, it can be speculated that both genes are engaged in DHA biosynthesis and DHA in Aurantiochytrium sp. could be produced by jointed actions of both FAS and PKS systems.

Fast Focal Point Correction in Prism-Coupled Total Internal Reflection Scanning Imager Using an Electronically Tunable Lens.

Total internal reflection (TIR) is useful for interrogating physical and chemical processes that occur at the interface between two transparent media. Yet prism-coupled TIR imaging microscopes suffer from limited sensing areas due to the fact that the interface (the object plane) is not perpendicular to the optical axis of the microscope. In this paper, we show that an electrically tunable lens can be used to rapidly and reproducibly correct the focal length of an oblique-incidence scanning microscope (OI-RD) in a prism-coupled TIR geometry. We demonstrate the performance of such a correction by acquiring an image of a protein microarray over a scan area of 4 cm² with an effective resolution of less than 20 microns. The electronic focal length tuning eliminates the mechanical movement of the illumination lens in the scanning microscope and in turn the noise and background drift associated with the motion.

Self-regulated viscous channel in the nuclear pore complex.

The nuclear pore complex (NPC), the sole gateway for nucleocytoplasmic exchange in eukaryotic cells, allows for the passive diffusion of small molecules and transport-receptor-facilitated translocation of signal-dependent cargo molecules. Whether small molecules passively diffuse through a single central channel or through multiple holes of a hydrogel network is a subject of debate. Additionally, whether the passive and facilitated transport systems occupy distinct or overlapping physical regions of the NPC remains unclear. Here, we directly test these models using three-dimensional super-resolution fluorescence microscopy of human cells. This approach reveals that a single viscous central channel in the NPC acts as the sole pathway for passive diffusion of various small molecules; transport receptors and their cargo complexes take distinct transport routes in the periphery, which is occluded by phenylalanine-glycine filaments. Furthermore, the passive and facilitated passageways in the NPC are closely correlated, and their conformations can be simultaneously regulated by Importin ß1 (a major transport receptor) and RanGTP (a critical regulator of transport directionality). These results strongly favor a self-regulated viscous channel configuration in native NPCs over the porous hydrogel meshwork model.

Genetic structure of duckweed population of Spirodela, Landoltia and Lemna from Lake Tai, China.

Duckweed is widely used in environmental biotechnology and has recently emerged as a potential feedstock for biofuels due to its high growth rate and starch content. The genetic diversity and composition of a natural duckweed population in genera Spirodela, Landoltia and Lemna from Lake Tai, China, were investigated using probabilistic analysis of multilocus sequence typing (MLST). The 78 strains were categorized into five lineages, among which strains representing L. aequinoctialis and S. polyrhiza were predominant. Among the five lineages, interlineage transfers of markers were infrequent and no recombination was statistically detected. Tajima's D tests determined that all loci are subject to population bottlenecks, which is likely one of the main reasons for the low genetic diversity observed within the lineages. Interestingly, strains of L. turionifera are found to contain small admixture from L. minor, providing rare evidence of transfer of genetic materials in duckweed. This was discussed with respect to the hypothesis that a cross of these two gave rise to L. japonica. Moreover, the conventional maximum-likelihood phylogenetic analysis clearly recognized all the species in the three genera with high bootstrap supports. In conclusion, this work offers a basic framework for using MLST to characterize Spirodela, Landoltia and in particular Lemna strains at the species level, and to study population genetics and evolution history of natural duckweed populations.

Ojeok-san enhances platinum sensitivity in ovarian cancer by regulating adipocyte paracrine IGF1 secretion.

BACKGROUND: Platinum is a commonly used drug for ovarian cancer (OvCa) treatment, but drug resistance limits its clinical application. This study intended to delineate the effects of adipocytes on platinum resistance in OvCa. METHODS: OvCa cells were maintained in the adipocyte-conditioned medium. Cell viability and apoptosis were detected by CCK-8 and flow cytometry, separately. Proliferation and apoptosis-related protein expression were assayed by western blot. The IC50 values of cisplatin and carboplatin were determined using CCK-8. IGF1 secretion and expression were assayed via ELISA and western blot, respectively. A xenograft model was established, and pathological changes were detected by H&E staining. Proliferation and apoptosis-associated protein expression was assessed via IHC. RESULTS: Adipocytes promoted the viability and repressed cell apoptosis in OvCa, as well as enhancing platinum resistance, while the addition of IGF-1 R inhibitor reversed the effects of adipocytes on proliferation, apoptosis, and drug resistance of OvCa cells. Treatment with different concentrations of Ojeok-san (OJS) inhibited the adipocyte-induced platinum resistance in OvCa cells by suppressing IGF1. The combined treatment of OJS and cisplatin significantly inhibited tumour growth in vivo with good mouse tolerance. CONCLUSION: In summary, OJS inhibited OvCa proliferation and platinum resistance by suppressing adipocyte paracrine IGF1 secretion.

Research and implementation of multi-disease diagnosis on chest X-ray based on vision transformer.

Background: Disease diagnosis in chest X-ray images has predominantly relied on convolutional neural networks (CNNs). However, Vision Transformer (ViT) offers several advantages over CNNs, as it excels at capturing long-term dependencies, exploring correlations, and extracting features with richer semantic information. Methods: We adapted ViT for chest X-ray image analysis by making the following three key improvements: (I) employing a sliding window approach in the image sequence feature extraction module to divide the input image into blocks to identify small and difficult-to-detect lesion areas; (II) introducing an attention region selection module in the encoder layer of the ViT model to enhance the model's ability to focus on relevant regions; and (III) constructing a parallel patient metadata feature extraction network on top of the image feature extraction network to integrate multi-modal input data, enabling the model to synergistically learn and expand image-semantic information. Results: The experimental results showed the effectiveness of our proposed model, which had an average area under the curve value of 0.831 in diagnosing 14 common chest diseases. The metadata feature network module effectively integrated patient metadata, further enhancing the model's accuracy in diagnosis. Our ViT-based model had a sensitivity of 0.863, a specificity of 0.821, and an accuracy of 0.834 in diagnosing these common chest diseases. Conclusions: Our model has good general applicability and shows promise in chest X-ray image analysis, effectively integrating patient metadata and enhancing diagnostic capabilities.

Li-Hong Tang alleviates dextran sodium sulfate-induced colitis by regulating NRF2/HO-1 signaling pathway and gut microbiota.

Introduction: Ulcerative colitis (UC) is marked by recurring inflammation. Existing treatments are ineffective and may have toxic side effects. Thus, new therapeutic agents are urgently needed. We studied the botanical formula "Li-Hong Tang (LHT)", which contains two main ingredients, Salvia plebeia R. Br and Rhodiola crenulata (Hook. f. et Thoms.) H. Ohba. In this study, we aimed to identify the effects of LHT on UC and explore its potential mechanism. Methods: LHT was analyzed using a mass spectrometer (MS). DSS at a dose of 2.5% was utilized to develop UC in mice. The administered groups received low, medium, and high dosages (0.32 g/kg, 0.64 g/kg, and 1.28 g/kg) of LHT and the positive medication, sulfasalazine (0.2 g/kg), respectively. Body weight, disease activity index (DAI) score, colon length, spleen index, serum myeloperoxidase (MPO), nitric oxide (NO), superoxide dismutase (SOD) and inflammatory factor concentrations were monitored. The expression of NRF2 and HO-1 in colonic tissues was evaluated by immunohistochemistry. 16S rDNA sequencing was employed to investigate alterations in the gut microbiota of the mice, aiming to elucidate the extent of LHT's impact. Results: LHT may ameliorate DSS-induced colitis in mice by lowering inflammation, reducing oxidative stress, restoring the intestinal barrier, and influencing the NRF2/HO-1 pathway. Moreover, LHT treatment exhibited a regulatory effect on the gut microbiota, characterized by elevated levels of Patescibacteria, Verrucomicrobiota, Candidatus_Saccharimonas, Lactobacillus, and Ligilactobacillus levels while decreasing Oscillibacter and Colidextribacter levels. Further study indicated that MPO, NO, and inflammatory factors were positively correlated with Oscillibacter, Colidextribacter, Escherichia-Shigella, Anaerostines, and negatively with Lactobacillus, Clostridiales_unclassified, Candidatus_Saccharimonas, and Patescibacteria. Furthermore, colony network analysis revealed that Lactobacillus was negatively associated with Oscillibacter and Colidextribacter, whereas Oscillibacter was positively related to Colidextribacter. Conclusion: LHT protects against DSS-induced mice by inhibiting the inflammatory response, oxidative stress, and mucosal injury. The protective role may involve regulating the NRF2/HO-1 signaling pathway and gut microbiota.

Enhancing the photodynamic effect of curcumin through modification with TiO2 nanoparticles and cationic polymers.

Curcumin (CUR), a natural compound extracted from turmeric, has shown potential as a photosensitizer in photodynamic therapy (PDT). The aim of this work was to enhance the efficacy of CUR by modifying it using titanium dioxide (TiO2) nanoparticles and a cationic polymer called Sofast to create a nanocomposite TiO2-CUR-Sofast (TCS). Compared to unmodified CUR, TCS exhibited a broadening toward longer wavelength in the absorption wavelength within the 400-550 nm range, leading to improved CUR absorption. Cellular uptake efficiency of TCS was also enhanced, and it demonstrated nearly 4.7-fold higher reactive oxygen species (ROS) generation than CUR. Furthermore, TCS displayed the ability to attach to the cell membrane and enter cells within a 30-min incubation period. Upon irradiation, TCS exhibited remarkable cytotoxicity, resulting in a significant reduction in the viability of various cancer cells. Autofluorescence lifetime imaging of intracellular reduced nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) and flavin adenine dinucleotide (FAD) enzymes indicated that cancer cells treated with TCS and irradiation undergo a metabolic pathway shift from oxidative phosphorylation to glycolysis. These findings highlight the potential of TCS as an effective PDT agent for cancer treatment.

Non-invasive screening of bladder cancer using digital microfluidics and FLIM technology combined with deep learning.

Non-invasive screening for bladder cancer is crucial for treatment and postoperative follow-up. This study combines digital microfluidics (DMF) technology with fluorescence lifetime imaging microscopy (FLIM) for urine analysis and introduces a novel non-invasive bladder cancer screening technique. Initially, the DMF was utilized to perform preliminary screening and enrichment of urine exfoliated cells from 54 participants, followed by cell staining and FLIM analysis to assess the viscosity of the intracellular microenvironment. Subsequently, a deep learning residual convolutional neural network was employed to automatically classify FLIM images, achieving a three-class prediction of high-risk (malignant), low-risk (benign), and minimal risk (normal) categories. The results demonstrated a high consistency with pathological diagnosis, with an accuracy of 91% and a precision of 93%. Notably, the method is sensitive for both high-grade and low-grade bladder cancer cases. This highly accurate non-invasive screening method presents a promising approach for bladder cancer screening with significant clinical application potential.

Three-dimensional distribution of transient interactions in the nuclear pore complex obtained from single-molecule snapshots.

The translocation of large macromolecules through the nuclear pore complex (NPC) of eukaryotic cells is hindered by the phenylalanine-glycine (FG) nucleoporin (Nup) barrier unless molecules are chaperoned by transport receptors. The precise mechanism of facilitated translocation remains unclear due to the challenges of measuring the series of transient interactions between a transport receptor and the FG-Nups. This study developed single-point edge-excitation subdiffraction microscopy to obtain a three-dimensional density map of the transient interactions with a spatiotemporal resolution of 9 nm and 400 mus. Three unique features were observed under real-time trafficking conditions that have escaped detection by conventional electron microscopy: (i) the spatial density of interaction sites between Importin beta1 (Imp beta1, a major transport receptor) and the FG-Nups gradually increases from both sides of the NPC and is highest in the central pore region; (ii) cargo-free or cargo-bound Imp beta1 rarely occupies an axial channel with a diameter of approximately 10-20 nm at its narrowest point through the NPC; and (iii) the pathway of facilitated translocation through the NPC depends more on the interaction sites of the FG-Nups than on the NPC architecture.