As a new kind of advanced nonlinear imaging approach, two-photon fluorescence microscopy technology is wildly used in the field of live cell and tissue imaging, especially focusing on long-term dynamic three-dimensional cell imaging. This paper firstly presents the principle and characteristic of two-photon fluorescence microscopy. Then, the paper focuses on the three key aspects of the viability of the specimen, sensitivity of detection, as well as the speed of acquisition. In the end, the future prospect of development and application of two-phonon imaging technology are predicted.
Diagnostic Imaging ; Microscopy, Fluorescence, Multiphoton ; methods

We applied two classical algorithms in hyperspectral imaging to extract endmember for biological fluorescence imaging. A combined algorithm was found to initialize the PPI with decreasing the number of multi-spectral pixels, with subsequent N-FINDR refinement, which could make a better result.
Algorithms ; Biology ; methods ; Diagnostic Imaging ; methods ; Image Interpretation, Computer-Assisted ; methods ; Microscopy, Fluorescence ; methods ; Spectrometry, Fluorescence ; methods

Animals ; Fluorescence ; Humans ; Microscopy, Fluorescence ; Nanotechnology ; Neoplasms ; diagnosis ; Photons ; Quantum Dots ; Staining and Labeling ; instrumentation ; methods

Confocal scanning microscopy is a new technique which can obtain the 3-D images of samples by using scanning and computer processing. It has been widely used in biomedical fields owing to its advantages such as high resolution and 3-D imaging. In this paper, we analyze the principle and research development of confocal scanning microscopies, and illustrate its advantages by comparing it with other microscopies.
Image Enhancement ; methods ; Image Processing, Computer-Assisted ; Imaging, Three-Dimensional ; methods ; Microscopy, Confocal ; methods ; Microscopy, Fluorescence ; methods ; Reproducibility of Results

OBJECTIVES: We produced hematoxylin and eosin (H&E) staining-like color images by using confocal laser scanning microscopy (CLSM), which can obtain the same or more information in comparison to conventional tissue staining. METHODS: We improved images by using several image converting techniques, including morphological methods, color space conversion methods, and segmentation methods. RESULTS: An image obtained after image processing showed coloring very similar to that in images produced by H&E staining, and it is advantageous to conduct analysis through fluorescent dye imaging and microscopy rather than analysis based on single microscopic imaging. CONCLUSIONS: The colors used in CLSM are different from those seen in H&E staining, which is the method most widely used for pathologic diagnosis and is familiar to pathologists. Computer technology can facilitate the conversion of images by CLSM to be very similar to H&E staining images. We believe that the technique used in this study has great potential for application in clinical tissue analysis.
Diagnosis ; Eosine Yellowish-(YS) ; Fluorescence ; Hematoxylin* ; Image Processing, Computer-Assisted ; Methods* ; Microscopy ; Microscopy, Confocal* ; Staining and Labeling

OBJECTIVETo investigate the microscopic autofluorescent characteristics of cardiac cancer and autofluorescence distribution in different layers of gastric tissues.

METHODSA double-channel laser scanning confocal microscopy with Argon ion laser (excitation wavelength 488 nm) and Helium-Neon laser (excitation wavelength 543 nm) were used to detect the autofluorescence emitted from 16 surgical specimens of cardiac cancer and corresponding normal gastric tissue. The autofluorescence image was analyzed between the cardiac cancer tissue and normal gastric tissue.

RESULTSAutofluorescence was detected successfully in cardiac carcinoma and corresponding normal gastric corpus tissues of all 16 cases. In different layers of gastric tissue, fluorescence presented the strongest signal in submucosa,the second strong in luminal propria with fluorescence mostly distributed in the glands, fluorescence signal from gastric cancer was significantly decreased compared with those in the different layers of normal tissues (P< 0.01).

CONCLUSIONThere are significant differences in the shape, color, distribution and fluorescence intensity of microscopic autofluorescence between cardiac cancer tissues and normal gastric corpus tissues.

Aged ; Female ; Heart Neoplasms ; pathology ; Humans ; Male ; Microscopy, Confocal ; Microscopy, Fluorescence ; methods ; Middle Aged ; Stomach ; pathology

In this paper, GLUT4 vesicles are observed in real-time under TIRF microscopy and a new three-dimensional single particle tracking algorithm according to the unique features of TIRF is put forward. Firstly a fluorescence correction procedure was processed to solve the problem of fluorescence bleaching over time and mobile vesicles were segmented by an adaptive background subtraction method. Kalman filtering was then introduced to track the granules so as to reduce the searching range and to avoid the disturbance of background noise and false targets. In the experiments the algorithm was applied in analyzing the long-distance movement of GLUT4 vesicles. The experimental results indicate that the algorithm has achieved robust tracking of the vesicles in the imaging plane and has effectively calculated the position in the direction orthogonal to the imaging plane.
Glucose Transporter Type 4 ; metabolism ; Imaging, Three-Dimensional ; instrumentation ; methods ; Ion Transport ; Microscopy, Fluorescence ; methods

Enzymes play such a pivotal role in cellular metabolism that enzyme assays are important for bio-engineering, disease diagnoses and drug discovery. Among the reported methods, fluoremetry has attracted more and more attention due to its high sensitivity and possibility of continuous dynamic monitoring. The recent progresses and applications in enzyme assays using fluorescent probes were reviewed. Different methods were classified into direct fluorescence detection and indirect fluorescence detection according to their labeled substrates and detection mechanisms. Our writing purpose is to provide the readers with a flavor of the kinds of tools and strategies available in enzyme assays with fluorescent probes. Also, the research situation and prospects were disucssed
Animals ; Enzyme Assays ; methods ; trends ; Fluorescent Dyes ; Fluorometry ; methods ; Humans ; Microscopy, Fluorescence

Quantitative analysis of biological image data generally involves the detection of many pixel spots. In live mitochondria video image, for which fluorescent microscopy is often used, the signal-to-noise ratio (SNR) can be extremely low, making the detection and tracking of mitochondria particle difficult. It is especially not easy to get the movement curve when the movement of the mitochondria involves its self-move and the motion caused by the neuron. An tracking algorithm for live mitochondria is proposed in this paper. First the whole image sequence is frame-to-frame registered, in which the edge corners are chosen to be the feature points. Then the mitochondria particles are tracked by frame-to-frame displacement vector. The algorithm proposed has been applied to the dynamic image sequence including neuron and mitochondria, saving time without manually picking up the feature points. It provides an new method and reference for medical image processing and biotechnological research.
Algorithms ; Animals ; Image Processing, Computer-Assisted ; methods ; Microscopy, Fluorescence ; Mitochondria ; metabolism ; ultrastructure ; Neurons ; ultrastructure ; Particle Size

Single molecule detection is a technology of studying biomolecules with high spatial and temporal resolution. By exploiting recent technical advances, we are able to observe, detect, even manipulate individual molecules and study their conformational changes and dynamic behaviors. New information can be obtained from the single molecule research, which is otherwise hidden or averaged out. In recent years, the development of single molecule detection techniques has opened up a new era of life science. In this review, we introduce the advances of the techniques for detecting single molecules in cell biology and review the development of single molecule detection in living cells.
Animals ; Biotechnology ; trends ; Cell Physiological Phenomena ; Humans ; Microscopy, Confocal ; methods ; Microscopy, Fluorescence ; instrumentation ; methods ; Molecular Biology ; instrumentation ; methods ; Nanotechnology ; instrumentation ; methods ; Proteins ; analysis ; ultrastructure