Chemical & Biomedical Imaging最新文献

The visualization of the central nervous system (CNS) has proposed stringent criteria for fluorescent probes, as the inevitable production of reactive oxygen species (ROS) or heat generated from most photoluminescent probes upon excitation can disturb the normal status of relatively delicate CNS cells. In this work, a red-emitting fluorogen with aggregation-induced emission (AIE) characteristics, known as DTF, was chosen as the model fluorogen to investigate whether the side effects of ROS and heat could be suppressed through easy-to-operate processes. Specifically, DTF was encapsulated with different amphiphilic matrices to yield AIE nanoprobes, and their photoluminescent properties, ROS production, and photothermal conversion rates were examined. BSA@DTF NPs possessed 1.3-fold brightness compared to that of DSPE-PEG@DTF NPs and F127@DTF NPs but its ROS generation efficiency is markedly decreased to only 2.4% of that produced by F127@DTF NPs. Meanwhile, BSA@DTF NPs showed a negligible photothermal effect. These features make BSA@DTF NPs favorable for long-term live cell imaging, particularly for fluorescent imaging of CNS cells. BSA@DTF NPs were able to sustain the normal state of HT-22 neuronal cells with continuous illumination for at least 25 min, and they also preserved the cytoskeleton of microglia BV-2 cells as the untreated control group. This work represents a successful but easy-to-operate process to suppress the ROS generation of red-emissive AIEgen, and it highlights the importance of minimizing the ROS generation of the fluorescent probes, particularly in the application of long-term imaging of CNS cells.

The visualization of the central nervous system (CNS) has proposed stringent criteria for fluorescent probes, as the inevitable production of reactive oxygen species (ROS) or heat generated from most photoluminescent probes upon excitation can disturb the normal status of relatively delicate CNS cells. In this work, a red-emitting fluorogen with aggregation-induced emission (AIE) characteristics, known as DTF, was chosen as the model fluorogen to investigate whether the side effects of ROS and heat could be suppressed through easy-to-operate processes. Specifically, DTF was encapsulated with different amphiphilic matrices to yield AIE nanoprobes, and their photoluminescent properties, ROS production, and photothermal conversion rates were examined. BSA@DTF NPs possessed 1.3-fold brightness compared to that of DSPE-PEG@DTF NPs and F127@DTF NPs but its ROS generation efficiency is markedly decreased to only 2.4% of that produced by F127@DTF NPs. Meanwhile, BSA@DTF NPs showed a negligible photothermal effect. These features make BSA@DTF NPs favorable for long-term live cell imaging, particularly for fluorescent imaging of CNS cells. BSA@DTF NPs were able to sustain the normal state of HT-22 neuronal cells with continuous illumination for at least 25 min, and they also preserved the cytoskeleton of microglia BV-2 cells as the untreated control group. This work represents a successful but easy-to-operate process to suppress the ROS generation of red-emissive AIEgen, and it highlights the importance of minimizing the ROS generation of the fluorescent probes, particularly in the application of long-term imaging of CNS cells.

Super-resolution optical imaging overcomes the diffraction limit in light microscopy to enable the visualization of previously invisible molecular details within a sample. The realization of super-resolution imaging based on stimulated Raman scattering (SRS) microscopy represents a recent area of fruitful development that has been used to visualize cellular structures in three dimensions, with multiple spectroscopic colors at the nanometer scale. Several fundamental approaches to achieving super-resolution SRS imaging have been reported, including optical engineering strategies, expansion microscopy, deconvolution image analysis, and photoswitchable SRS reporters as methods to break the diffraction limit. These approaches have enabled the visualization of biological structures, cellular interactions, and dynamics with unprecedented detail. In this Perspective, an overview of the current strategies and capabilities for achieving super-resolution SRS imaging will be highlighted together with an outlook on potential directions of this rapidly evolving field.

Super-resolution optical imaging overcomes the diffraction limit in light microscopy to enable the visualization of previously invisible molecular details within a sample. The realization of super-resolution imaging based on stimulated Raman scattering (SRS) microscopy represents a recent area of fruitful development that has been used to visualize cellular structures in three dimensions, with multiple spectroscopic colors at the nanometer scale. Several fundamental approaches to achieving super-resolution SRS imaging have been reported, including optical engineering strategies, expansion microscopy, deconvolution image analysis, and photoswitchable SRS reporters as methods to break the diffraction limit. These approaches have enabled the visualization of biological structures, cellular interactions, and dynamics with unprecedented detail. In this Perspective, an overview of the current strategies and capabilities for achieving super-resolution SRS imaging will be highlighted together with an outlook on potential directions of this rapidly evolving field.

Studying embryogenesis is fundamental to understanding developmental biology and reproductive medicine. Its process requires precise spatiotemporal regulations in which lipid metabolism plays a crucial role. However, the spatial dynamics of lipid species at the subcellular level remains obscure due to technical limitations. To address this challenge, we developed a hyperspectral 3D imaging and analysis method based on stimulated Raman scattering microscopy (hyper-3D SRS) to quantitatively assess lipid profiles in individual embryos through submicrometer resolution (x–y), 3D optical sectioning (z), and chemical bond-selective (Ω) imaging. Using hyper-3D SRS, individual lipid droplets (LDs) in single cells were identified and quantified. Our findings revealed that the LD profiles within a single embryo are not uniform, even as early as the 2-cell stage. Notably, we also discovered a dynamic relationship between the LD size and unsaturation degree as embryos develop, indicating diverse lipid metabolism during early development. Furthermore, abnormal LDs were observed in oocytes of a progeria mouse model, suggesting that LDs could serve as a potential biomarker for assessing oocyte/embryo quality. Overall, our results highlight the potential of hyper-3D SRS as a noninvasive method for studying lipid content, composition, and subcellular distribution in embryos. This technique provides valuable insights into lipid metabolism during embryonic development and has the potential for clinical applications in evaluating oocyte/embryo quality.

Studying embryogenesis is fundamental to understanding developmental biology and reproductive medicine. Its process requires precise spatiotemporal regulations in which lipid metabolism plays a crucial role. However, the spatial dynamics of lipid species at the subcellular level remains obscure due to technical limitations. To address this challenge, we developed a hyperspectral 3D imaging and analysis method based on stimulated Raman scattering microscopy (hyper-3D SRS) to quantitatively assess lipid profiles in individual embryos through submicrometer resolution (x-y), 3D optical sectioning (z), and chemical bond-selective (Ω) imaging. Using hyper-3D SRS, individual lipid droplets (LDs) in single cells were identified and quantified. Our findings revealed that the LD profiles within a single embryo are not uniform, even as early as the 2-cell stage. Notably, we also discovered a dynamic relationship between the LD size and unsaturation degree as embryos develop, indicating diverse lipid metabolism during early development. Furthermore, abnormal LDs were observed in oocytes of a progeria mouse model, suggesting that LDs could serve as a potential biomarker for assessing oocyte/embryo quality. Overall, our results highlight the potential of hyper-3D SRS as a noninvasive method for studying lipid content, composition, and subcellular distribution in embryos. This technique provides valuable insights into lipid metabolism during embryonic development and has the potential for clinical applications in evaluating oocyte/embryo quality.

Macrophages exhibit a spectrum of behaviors upon activation and are generally classified as one of two types: inflammatory (M1) or anti-inflammatory (M2). Tracking these phenotypes in living cells can provide insight into immune function but remains a challenging pursuit. Existing methods are mostly limited to static readouts or are difficult to employ for multiplexed imaging in complex 3D environments while maintaining cellular resolution. We aimed to fill this void using bioluminescent technologies. Here we report genetically engineered luciferase reporters for the long-term monitoring of macrophage polarization via spectral phasor analysis. M1- and M2-specific promoters were used to drive the expression of bioluminescent enzymes in macrophage cell lines. The readouts were multiplexed and discernible in both 2D and 3D formats with single-cell resolution in living samples. Collectively, this work expands the toolbox of methods for monitoring macrophage polarization and provides a blueprint for monitoring other multifaceted networks in heterogeneous environments.

Macrophages exhibit a spectrum of behaviors upon activation and are generally classified as one of two types: inflammatory (M1) or anti-inflammatory (M2). Tracking these phenotypes in living cells can provide insight into immune function but remains a challenging pursuit. Existing methods are mostly limited to static readouts or are difficult to employ for multiplexed imaging in complex 3D environments while maintaining cellular resolution. We aimed to fill this void using bioluminescent technologies. Here we report genetically engineered luciferase reporters for the long-term monitoring of macrophage polarization via spectral phasor analysis. M1- and M2-specific promoters were used to drive the expression of bioluminescent enzymes in macrophage cell lines. The readouts were multiplexed and discernible in both 2D and 3D formats with single-cell resolution in living samples. Collectively, this work expands the toolbox of methods for monitoring macrophage polarization and provides a blueprint for monitoring other multifaceted networks in heterogeneous environments.

The large-scale preparation of fluorescent nanomaterials with laboratory-relevant chemical and optical properties will greatly forward their consumer market applications; however, it still remains challenging. In this work, a universal strategy was developed for the rapid and large-scale synthesis of fluorescent sulfur quantum dots that recently has drawn great attention because of their unique optical characteristics. From the fact that empty 3d orbitals of sulfide species are able to bind with lone-pair π electrons of the heteroatomic groups, many amino-group containing compounds, such as amino acid and polyethylenimine molecules, were exploited to synthesize sulfur quantum dots. This 10 min preparation period endowed sulfur quantum dots with bright blue fluorescence and also chirality. Due to the user-friendly and rapid operation, this strategy can be extended to the large-scale synthesis of sulfur quantum dots with a yield of 16.844 g for one batch of experiment. Moreover, it was found that the sulfur quantum dots exhibited a reversible temperature-dependent luminescent property with a sensitivity of 0.72%/°C, which showed excellent intracellular temperature monitoring capability for inflammation-related disease diagnostics.

The large-scale preparation of fluorescent nanomaterials with laboratory-relevant chemical and optical properties will greatly forward their consumer market applications; however, it still remains challenging. In this work, a universal strategy was developed for the rapid and large-scale synthesis of fluorescent sulfur quantum dots that recently has drawn great attention because of their unique optical characteristics. From the fact that empty 3d orbitals of sulfide species are able to bind with lone-pair π electrons of the heteroatomic groups, many amino-group containing compounds, such as amino acid and polyethylenimine molecules, were exploited to synthesize sulfur quantum dots. This 10 min preparation period endowed sulfur quantum dots with bright blue fluorescence and also chirality. Due to the user-friendly and rapid operation, this strategy can be extended to the large-scale synthesis of sulfur quantum dots with a yield of 16.844 g for one batch of experiment. Moreover, it was found that the sulfur quantum dots exhibited a reversible temperature-dependent luminescent property with a sensitivity of 0.72%/°C, which showed excellent intracellular temperature monitoring capability for inflammation-related disease diagnostics.