Chemical & Biomedical Imaging最新文献

An uncontrolled immune response leads to many diseases; therefore, monitoring inflammation is crucial for the diagnosis of subsequent diseases, drug screening, and targeted therapy. Since the inflammatory response mainly occurs in macrophages, there is a need to develop more inflammatory probes with macrophage-targeting ability. Herein, we designed a macrophage-targeted and hydrogen-peroxide-activated fluorescent probe BOH-HCy-Man for real-time imaging of inflammation in vivo and a control probe BOH-HCy without the macrophage-targeting part. The larger rate constant toward H₂O₂ led to the higher sensitivity of BOH-HCy-Man (19.1-fold) than BOH-HCy (10.2-fold) in vitro. With the help of its macrophage-targeting ability, BOH-HCy-Man possessed an additional 1.6-fold fluorescent enhancement in inflamed RAW 264.7 cells or 1.3-fold fluorescent enhancement in vivo than BOH-HCy. We expected that BOH-HCy-Man will be a powerful tool for early diagnosis of inflammation related diseases.

Lipid rafts (LRs) are relatively well-ordered functional microdomains in cell membranes and play an irreplaceable role in physiological processes as a transduction platform for multiple signaling pathways. Due to their small size and high spatiotemporal dynamics, it is difficult to perform lipid raft-localized biomolecule imaging on the surface of living cells. Here, we report a DNA nanotechnology-based platform for reversible manipulation and localized analysis of lipid rafts, which consists of two modules: “patching and coding probe pair” and “fishing probe”. The probe pair is generated by modifying two different sets of connectable DNA structures on a lipid raft-specific protein. After recognizing lipid rafts, the two probes in close proximity are linked by a DNA ligase reaction to form a lipid raft identity (LR-ID) code. The LR-ID strand patches and stabilizes the lipid raft structure. Interestingly, the raft patches formed can be depatched by restriction endonucleases, providing the first reversible manipulation of the lipid raft structure in living cells. We also designed a “fishing probe” with a DNA hairpin structure using an aptamer that can specifically bind to the target. The probe can cascade the reaction to two input signals “LR-ID” and “target protein” to generate an “off–on” fluorescence switch, allowing imaging and dynamic monitoring of target proteins localized in lipid rafts. By encoding arbitrary targets (in the case of glycans) in lipid rafts, we have created a universal lipid raft-localized imaging platform. This work provides an integrated analytical and manipulative platform to reveal lipid rafts and associated signaling pathways at the molecular level.

Brachytherapy is an established treatment modality that has been globally utilized for the therapy of malignant solid tumors. However, classic therapeutic sealed sources used in brachytherapy must be surgically implanted directly into the tumor site and removed after the requisite period of treatment. In order to avoid the trauma involved in the surgical procedures and prevent undesirable radioactive distribution at the cancerous site, well-dispersed radiolabeled nanomaterials are now being explored for brachytherapy applications. This emerging field has been coined “nanoscale brachytherapy”. Despite present-day advancements, an ongoing challenge is obtaining an advanced, functional nanomaterial that concurrently incorporates features of high radiolabeling yield, short labeling time, good radiolabeling stability, and long tumor retention time without leakage of radioactivity to the nontargeted organs. Further, attachment of suitable targeting ligands to the nanoplatforms would widen the nanoscale brachytherapy approach to tumors expressing various phenotypes. Molecular imaging using radiolabeled nanoplatforms enables noninvasive visualization of cellular functions and biological processes in vivo. In vivo imaging also aids in visualizing the localization and retention of the radiolabeled nanoplatforms at the tumor site for the requisite time period to render safe and effective therapy. Herein, we review the advancements over the last several years in the synthesis and use of functionalized radiolabeled nanoplatforms as a noninvasive substitute to standard brachytherapy sources. The limitations of present-day brachytherapy sealed sources are analyzed, while highlighting the advantages of using radiolabeled nanoparticles (NPs) for this purpose. The recent progress in the development of different radiolabeling methods, delivery techniques and nanoparticle internalization mechanisms are discussed. The preclinical studies performed to date are summarized with an emphasis on the current challenges toward the future translation of nanoscale brachytherapy in routine clinical practices.

Vomocytosis is a process that occurs when internalized fungal pathogens escape from phagocytes without compromising the viability of the pathogen and the host cell. Manual quantification of time-lapse microscopy videos is currently used as the standard to study pathogen behavior and vomocytosis incidence. However, human-driven quantification of vomocytosis (and the closely related phenomenon, exocytosis) is incredibly burdensome, especially when a large volume of cells and interactions needs to be analyzed. In this study, we designed a MATLAB algorithm that measures the extent of colocalization between the phagocyte and fungal cell (Cryptococcus neoformans; CN) and rapidly reports the occurrence of vomocytosis in a high throughput manner. Our code processes multichannel, time-lapse microscopy videos of cocultured CN and immune cells that have each been fluorescently stained with unique dyes and provides quantitative readouts of the spatiotemporally dynamic process that is vomocytosis. This study also explored metrics, such as the rate of change of pathogen colocalization with the host cell, that could potentially be used to predict vomocytosis occurrence based on the quantitative data collected. Ultimately, the algorithm quantifies vomocytosis events and reduces the time for video analysis from over 1 h to just 10 min, a reduction in labor of 83%, while simultaneously minimizing human error. This tool significantly minimizes the vomocytosis analysis pipeline, accelerates our ability to elucidate unstudied aspects of this phenomenon, and expedites our ability to characterize CN strains for the study of their epidemiology and virulence.

With the flourishing development of precision medicine, theranostics, generally recognized as the integration of diagnosis and treatment, has emerged as a prominent trend in clinical research. However, theranostics primarily emphasizes the end result of integration, without providing sufficient details on how precise diagnosis and synergetic individualized treatment could be achieved and what clinical challenges could be effectively addressed in clinical practice. Molecular probe technology provides a robust method to bridge the gap between theory and practice. Through meticulous design of the chemical structure, imaging labels or drugs were conjugated to tumor-targeting peptides, antibodies, or inducers to form molecular probes, which allow a seamless switch between targeted intervention and targeted imaging with consistency in time, space, and biodistribution. Thus, this review proposes a concept called “molecular eye”, which refers to a comprehensive system for precise diagnosis and treatment of major clinical diseases based on molecular probe technology. This medical system emphasizes the chemical basis of probe development and optimization, which can provide precise actionable information for clinical decision making, allow molecular-targeted therapy, expand the indications of old therapy, and accelerate the regulatory approval of molecular drugs. “Molecular eye” resembles the piercing eye of the Monkey King, which can detect previously “invisible” diseases and facilitate disease diagnosis, treatment, real-time evaluation, and pathology research, guiding drug development. The emergence of the “molecular eyes” will provide opportunities and challenges in the fields of clinical practice and medical research and propel the progression of contemporary medicine toward precision medicine.

Optical clearing of invertebrates, the number of species of which is 20 times greater than that of vertebrates, is of fundamental and applied interest for neuroscience in general. Herein, the optical clearing of invertebrates to identify their morphology and neurostructure remains unrealized as of yet. Here, we report on fast (from a few seconds to minutes) and uniform whole-body optical clearing of invertebrates (bivalves, nemertines, annelids, and anomura) of any age and thickness (up to 2 cm) possessing complicated structures and integuments. We developed the protocol unifying dimethyl sulfoxide (DMSO)-based immunostaining of the animals followed by their optical clearing with benzyl alcohol/benzyl benzoate (BABB). Confocal microspectroscopy revealed that the protocol provides an increase of the fluorescence signal by 2 orders of magnitude and decrease of the light scattering by 2 orders of magnitude, thereby accelerating the confocal bioimaging of the whole body. Moreover, by tracking the optical clearing over time with 0.3 s resolution, we revealed that the clearing process is described by the Gompertz growth function, allowing us to determine the physical mechanism of the clearing and its optical parameters. Thereby, we were able to identify in detail and to describe previously unknown neurostructures of different invertebrate animals, paving the way to discovery in neuroscience.

Gold nanoparticles are frequently employed as nanozyme materials due to their capacity to catalyze various enzymatic reactions. Given their plasmonic nature, gold nanoparticles have also found extensive utility in chemical and photochemical catalysis owing to their ability to generate excitons upon exposure to light. However, their potential for plasmon-assisted catalytic enhancement as nanozymes has remained largely unexplored due to the inherent challenge of rapid charge recombination. In this study, we have developed a strategy involving the encapsulation of gold nanorods (AuNRs) within a titanium dioxide (TiO₂) shell to facilitate the efficient separation of hot electron/hole pairs, thereby enhancing nanozyme reactivity. Our investigations have revealed a remarkable 10-fold enhancement in reactivity when subjected to 530 nm light excitation following the introduction of a TiO₂ shell. Leveraging single-molecule kinetic analyses, we discovered that the presence of the TiO₂ shell not only amplifies catalytic reactivity by prolonging charge relaxation times but also engenders additional reactive sites within the nanozyme’s intricate structure. We anticipate that further enhancements in nanozyme performance can be achieved by optimizing interfacial interactions between plasmonic metals and semiconductors.

Red blood cell (RBC) indices serve as clinically important parameters for diagnosing various blood-related diseases. Conventional hematology analyzers provide the highly accurate detection of RBC indices but require large blood volumes (>1 mL), and the results are bulk mean values averaged over a large number of RBCs. Moreover, they do not provide quantitative information related to the morphological and chemical alteration of RBCs at the single-cell level. Recently, quantitative phase imaging (QPI) methods have been introduced as viable detection platforms for RBC indices. However, coherent QPI methods are built on complex optical setups and suffer from coherent speckle noise, which limits their detection accuracy and precision. Here, we present spectroscopic differential phase-contrast (sDPC) microscopy as a platform for measuring RBC indices. sDPC is a computational microscope that produces color-dependent phase images with higher spatial resolution and reduced speckle noise compared to coherent QPIs. Using these spectroscopic phase images and computational algorithms, RBC indices can be extracted with high accuracy. We experimentally demonstrate that sDPC enables the high-accuracy measurement of the mean corpuscular hemoglobin concentration, mean corpuscular volume, mean corpuscular hemoglobin, red cell distribution width, hematocrit, hemoglobin concentration, and RBC count with errors smaller than 7% as compared to a clinical hematology analyzer based on flow cytometry (XN-2000; Sysmex, Kobe, Japan). We further validate the clinical utility of the sDPC method by measuring and comparing the RBC indices of the control and anemic groups against those obtained using the clinical hematology analyzer.

Wide-field photobleaching imprinting microscopy (PIM) can improve fluorescence image contrast by cleverly exploiting the fluorophores’ photobleaching properties. However, as conventional wide-field PIM commonly adopts Gaussian illumination with a nonuniform lateral fluence distribution, the field-of-view (FOV) and sampling density are largely reduced. In addition, the slow axial fluence gradient of Gaussian illumination limits the signal-to-background ratio (SBR) improvement and optical sectioning capability of PIM. Here, we present flat-field photobleaching imprinting microscopy (ffPIM) with a uniform lateral excitation fluence and sharp axial intensity gradient at the focal plane. ffPIM demonstrates low background, large FOV, and thin optical section. More importantly, compared to either conventional wide-field PIM or light-sheet microscopy, ffPIM shows much better balance for FOV, sampling density, SBR, and optical sectioning capability. The performance of ffPIM is characterized by simulation and resolving multiple cellular structures. Finally, ffPIM can be easily implemented to a standard commercial wide-field microscope and, thereby, allow general laboratories to benefit from this technique.