Unlike ensemble-averaging measurements, single-molecule tracking provides quantitative information on the kinetics of individual molecules within living cells in real time and may provide insight into the respective molecular interactions behind that. The advancement of single-molecule tracking has been significantly boosted by the development of high-resolution microscopy techniques. In this review, we will discuss this aspect with a particular focus on their recent advance in MINFLUX nanoscopy with feedback approaches where tracking is performed in real time. MINFLUX localization requires fewer than 100 photons from a ～1 nm-sized fluorophore, enabling precise tracking. This approach, which demands over an order of magnitude fewer photons than other localization-based techniques (such as STORM, PLAM), allows molecular tracking with single-digit nanometer accuracy in less than 1ms — an achievement previously unattainable.

Accurate identification and viability assessment of the parathyroid glands (PGs) are critical when performing thyroid and parathyroid surgeries. Traditional visual inspection-based intraoperative methods suffer from subjectivity and uncertainty. Near-infrared (NIR) imaging methods, including NIR autofluorescence (NIRAF) imaging and indocyanine green fluorescence imaging (ICGFI), have emerged as promising and reliable techniques for intraoperative PG identification and assessment. Here, the principles and clinical performance of NIR imaging methods were comprehensively reviewed.

As an emerging powerful tool to provide structural information of tissue specimens label-freely, Mueller matrix (MM) polarimetry has garnered extensive attention in biomedical studies and pathological diagnosis. However, for the commonly used constant-step rotating MM polarimetric system, beam drift induced by the rotation of polarization elements can lead to distortions in measurement results, severely affecting MM imaging accuracy. Here, based on our previous study, we propose an optimized self-registration method to mitigate the pseudo-depolarization effects introduced by image artifacts in constant-step rotating MM polarimetry. By addressing the prevalent issue of beam drift and image distortions in such polarimetric imaging systems, the effectiveness of the proposed method is experimentally validated using tissue samples. The results demonstrate a significant enhancement in the accuracy of depolarization parameter estimation after applying the optimized self-registration method. Furthermore, the method enhances the coarseness and contrast of MM-derived parameters images, thereby bolstering their capacity to characterize tissue structures. The optimized self-registration method proposed in this study can provide an innovative approach for quantitative tissue polarimetry based on constant-step rotating MM measurement, and contribute to the advancement of polarimetric imaging technology in biomedical applications.

In clinical environments, the prolonged utilization of polarization equipment can result in the accumulation of errors over extended periods. The absence of expeditious calibration techniques in clinical practice presents a significant obstacle in preserving the precision and dependability of these instruments. To address this challenge, we propose an innovative research study that presents a comprehensive calibration system specifically designed for the calibration of the backscattering Mueller matrix measurement system, enabling swift online calibration across various scenarios. This system employs an external calibration framework for real-time adjustment of the polarizer’s initial angle, overseeing the rotation of PSG and PSA motors through position measurement and control procedures, with light intensity monitored by a camera. By incorporating momentum concepts and the Adam optimization algorithm, we enhance convergence speed, mitigate noise, and improve calibration accuracy. Experimental results showcase the exceptional precision, speed, and robustness of our proposed method, achieving high accuracy and minimal error, thereby offering a promising solution for maintaining the reliability of polarization equipment in clinical settings.

Mueller matrix polarimetry (MMP) has been proven to be a powerful tool for characterizing the microstructural features of biological samples in biomedical research and clinical diagnostics. However, the traditional Mueller matrix (MM) imaging technique based on single exposure has a limited dynamic range, leading to poor polarization image quality for biological samples with significant contrast variations. In this study, we propose a novel method to generate high dynamic range (HDR) MM images based on a multi-exposure fusion algorithm. By employing an optimal exposure selection strategy for transmission imaging and a multi-exposure weighted averaging strategy for backscattering imaging, the method expands the dynamic range while accurately preserving the polarization information of the samples. Experiments of sliced and bulk tissues demonstrate that the proposed method significantly suppresses the scattering noise and improves the quality of extracted polarization parameter images, especially in accurate distinction of different pathological areas. These results highlight the potential of HDR MM imaging technology in extracting polarization information from complex biological samples with high resolution and contrast.

Triple-negative breast cancer (TNBC) metastasis is particularly severe due to its aggressive nature, leading to rapid disease progression and significantly reduced survival rates. Rujifang (RJF), a traditional Chinese formula, has demonstrated potential anti-tumor effects and the ability to inhibit TNBC metastasis. However, the effects of varying RJF doses remain unclear. This study utilized laser-based in vivo flow cytometry (IVFC) to monitor circulating tumor cells (CTCs) and evaluate the efficacy of RJF at different doses. The results indicated that RJF at the high dose inhibited both the number of CTCs and the formation of metastatic foci more effectively compared to the lower dose. TUNEL assays revealed that RJF treatment promotes apoptosis of tumor cells, with a more pronounced effect observed at the higher dose. Immunofluorescence experiments demonstrated that administering a higher dose of RJF suppresses the expression of Kindlin-1 more effectively in the tumor microenvironment. Although higher doses showed enhanced efficacy, they might also lead to an increase in side effects. These findings underscore the promise and challenges of using RJF at high doses for anti-tumor therapy. They highlight the critical importance of optimizing the dose of RJF in the treatment of TNBC and provide valuable insights for its clinical application.

In recent years, depression has emerged as a significant global health concern, prompting many individuals to seek pharmacological interventions. The identification of inflammatory changes in the hippocampus of depressed patients has highlighted a potential therapeutic target. Nevertheless, the effectiveness of medications targeting these specific alterations has yet to be fully substantiated. Preliminary research has suggested the potential benefits of photobiomodulation (PBM) as a treatment for depression, with no significant adverse effects reported. This study utilized near-infrared light at intensities of 50mW/cm2 and 300mW/cm2 to illuminate mice with chronic mild stress (CMS)-induced depression model, aiming to explore the therapeutic effects of PBM on depression. The findings revealed that when exposed to a power density of 300mW/cm2, the mice exhibited enhanced behavioral outcomes, accompanied by decreased levels of inflammatory cytokines such as IL-1α, IL-1β, IL-5, and IL-6 in the hippocampus. A noteworthy association was observed between behavioral manifestations and inflammatory cytokine levels. This study posits that PBM at an intensity of 300mW/cm2 is a viable nonpharmacological intervention for depression, as it demonstrates a notable enhancement in depressive symptoms and the regulation of inflammatory mediators within the hippocampal region of the brain. However, this study is constrained by the particular PBM parameters employed; therefore, additional research is necessary to investigate a broader spectrum of doses and treatment durations in order to enhance the therapeutic application and deepen the understanding of the underlying mechanisms.

Our goal was to develop and experimentally validate a polarization-interference method for phase scanning of laser speckle fields generated by diffuse layers of birefringent biological tissues. This method isolates and uses new diagnostic parameters related to the “phase waves of local depolarization”. We combined polarization-interference registration with phase scanning of complex amplitude distributions in diffuse laser speckle fields to detect phase waves of local depolarization in birefringent fibrillar networks of biological tissue and measure their modulation depth. This approach led to the discovery of new criteria for differentiating various necrotic changes in diffuse histological samples of myocardial tissue from deceased individuals with “ischemic heart disease (IHD) — acute coronary insufficiency (ACI)”, even in the presence of a high level of depolarized background. To evaluate the degree of necrotic changes in the optical anisotropy of diffuse myocardial layers, a new quantitative parameter — modulation depth of local depolarization wave fluctuations — has been proposed. Using this approach, for the first time, differentiation of diffuse myocardial samples from deceased individuals with IHD and ACI was achieved with a very good 90.45% and outstanding accuracy of 95.2%.

Ultrasound neuromodulation is a powerful tool for brain investigation and holds great promise for treating brain diseases. However, due to the heterogeneous acoustic properties of skulls, existing ultrasound neuromodulation faces the challenge of severe transcranial acoustic attenuation. To overcome such limitations, we report an implantable bio-chip for visible and controllable microwave-induced transcranial acoustic generation (MI-tAG). The bio-chip is soft, flexible, and biocompatible, with a thickness of 3mm, making it suitable for human intracranial implantation. The constituted fluid channels can cover an area of 50mm × 60mm, enabling widefield neuron stimulation. The particles filled in the fluid channels have both high microwave absorption, ensuring efficient ultrasound generation, and magnetism, allowing noncontact and flexible manipulation by external magnetic fields. The experimental results demonstrate that the optimal MI-tAG can be realized by the combination of particles arranged in a linear pattern and corresponding illumination via a linearly polarized microwave. Stability evaluation indicates that the particles can maintain a consistent acoustic intensity without degradation for at least seven days. The results of in vitro and in vivo experiments show that the MI-tAG can manipulate ultrasound sources and visibly locate them in real time. This study provides a potential innovative approach for future ultrasound neuromodulation, inspiring the development of more useful methods to advance brain research. This study introduces a promising innovative approach for transcranial acoustic generation, potentially inspiring the development of more effective methods for advancing ultrasound neuromodulation.

Distinguishing the severity of burned skin from structural optical coherence tomography (OCT) intensity maps remains a challenging task, and functional imaging from an elastic perspective can improve the accuracy of burned skin examination. As a functional extension of OCT, optical coherence elastography (OCE) can reveal the mechanical properties of samples while inheriting the imaging advantages of OCT. In this study, we used OCE to reveal the shear modulus and anisotropy parameters of burned skin before and after burning. A porcine skin burn model was constructed at a series of burned time durations and tested by elastic anisotropy imaging. Normal skin after hydration maintains good consistency in shear modulus. Interestingly, the shear modulus and longitudinal modulus of the burned skin show a tendency to stepwise increase with increasing burned times. A dataset was constructed by sampling the modulus parameters of burned skin maps through a scratch window, and its category was automatically identified by K-means and density peak clustering (DPC) algorithms with good agreement. The elastic anisotropy-based skin burn assessment method shows a prospect to be supplemented into the nondestructive means of burned skin examination.

The brain lymphatic system plays a crucial role in maintaining homeostasis, clearing metabolic waste, and regulating neuroinflammation. Its dysfunction is strongly linked to neurodegenerative diseases such as Alzheimer’s disease and Parkinson’s disease. In this study, we employed dual-contrast functional photoacoustic microscopy to evaluate the impact of lipopolysaccharide-induced central nervous system inflammation on brain lymphatic function and to explore the protective effects of the P2X7 receptor (P2X7R) antagonist. Our findings demonstrated that lipopolysaccharide intervention led to impaired function of the meningeal lymphatic vessels, which was partially restored by the P2X7R antagonist, whereas its effects on the glymphatic system and cerebral vessels were minimal. This study further supports the feasibility of photoacoustic microscopy for assessing brain lymphatic function and highlights the therapeutic potential of P2X7R antagonism. These findings suggest that P2X7R may serve as a key target for modulating brain-lymphatic interactions, providing an experimental foundation for developing intervention strategies for neuroinflammatory and neurodegenerative diseases.

Candida albicans (C. albicans), a common pathogenic fungus in nature, has enough capacity to cause severe brain infection through various means under immunocompromised conditions. Currently, establishing a basic animal disease model has become the main research tool, which is conducive to simulating fungal encephalitis effectively. However, the widely used bloodborne infection model established by intravenous (I.V) injection in mice usually results in systemic infections but cannot simulate significant brain inflammation. Here, we developed a fungal encephalitis model by intracerebroventricular (I.C.V) injection of C. albicans to better simulate the significant harm and consequences. Compared with I.V, a greater number of colony-forming units (CFUs) in the brain was induced following I.C.V. Magnetic resonance imaging (MRI) results revealed more obvious inflammation in the external capsule area of the brain. Meanwhile, behavioral experiments with the Y-maze also indicated that abnormal activity behavior further reflected significant short-term memory impairment after I.C.V of C. albicans. In summary, these studies not only provide a novel fungal encephalitis model for understanding the pathogenesis mechanism of this disease but also lay a solid foundation for future effective treatment.