Deep learning method for cell count from transmitted-light microscope

Automatic cell counting provides an effective tool for medical research and diagnosis.Currently,cell counting can be completed by transmitted-light microscope,however,it requires expert knowledge and the counting accuracy which is unsatisfied for overlapped cells.Further,the image-translation-based detection method has been proposed and the potential has been shown to accomplish cell counting from transmitted-light microscope,automatically and effectively.In this work,a new deep-learning(DL)-based two-stage detection method(cGAN-YOLO)is designed to further enhance the performance of cell counting,which is achieved by combining a DL-based fluorescent image translation model and a DL-based cell detection model.The various results show that cGAN-YOLO can effectively detect and count some different types of cells from the acquired transmitted-light microscope images.Compared with the previously reported YOLO-based one-stage detection method,high recognition accuracy(RA)is achieved by the cGAN-YOLO method,with an improvement of 29.80%.Furthermore,we can also observe that cGAN-YOLO obtains an improvement of 12.11%in RA compared with the previously reported image-translation-based detection method.In a word,cGAN-YOLO makes it possible to implement cell counting directly from the experimental acquired transmitted-light microscopy images with high flexibility and performance,which extends the applicability in clinical research.

Development of a portable reflectance confocal microscope and its application in the noninvasive in vivo evaluation of mesenchymal stem cell-promoted cutaneous wound healing

The process of wound healing is routinely evaluated by histological evaluation in the clinic,which may cause scarring and secondary injury.Reflectance confocal microscopy(RCM)represents a noninvasive,real-time imaging technique that allows in vivo evaluation of the skin.Traditional RCM was wide-probe-based,which limited its application on uneven and covered skin.In this study,we report the development of a portable reflectance confocal microscope(PRCM)in which all components were assembled in a handheld shell.Although the size and weight of the PRCM were reduced based on the use of a microelectromechanical system,the resolution was kept at 0.91μm,and the field of view of the system was 343μm×532μm.When used in vivo,the PRCM was able to visualize cellular and nuclear morphology for both mouse and human skin.PRCM evaluations were then performed on wounds after topically applied mesenchymal stem cells(MSCs)or saline treatment.The PRCM allowed visualization of the formation of collagen bundles,re-epithelization from the wound edge to the wound bed,and hair follicle regeneration,which were consistent with histological evaluations.Therefore,we offer new insights into monitoring the effects of topically applied MSCs on the process of wound healing by using PRCM.This study illustrates that the newly developed PRCM represents a promising device for real-time,noninvasive monitoring of the dynamic process of wound healing,which demonstrates its potential to diagnose,monitor,or predict disease in clinical wound therapy.

Comparative Alpha Tracks Counting Using an Optical Microscope and a Spark Counter

In the metrology of radon, an environmental lung carcinogen, the integrated measurements necessary for epidemiological studies are made very often using the tracks detector LR 115 type 2. For dosimetric analysis, the etched tracks from radon alpha particles on this detector are usually counted by means of an optical microscope or a spark counter. An optimal reading of the track densities which must be converted into radon concentrations, can’t be done without a good mastery of the mode of operation and use of these devices. Furthermore, investigations to know as to whether or not each of those can be used to determine radon concentration are necessary. These are the objectives of the present work in which LR 115 samples exposed to radon for at least 3 months, were chemically developed under standard conditions and read. The track densities obtained with the microscope are very much higher than those of the counter for each sample. These results are consistent with those published by other authors. However, each of these devices can be used interchangeably for alpha tracks counting, as both provide radon concentrations with a very good linear correlation coefficient of 0.95 taking into account their respective calibration factors for the reading of this detector. In addition, the saturation phenomenon for the spark counter reading of LR 115 detector occurs beyond 11,000 tr/cm2, a density never reached during our environmental radon measurements.

用IM Microscope方法计算无源互调产物

在微波与射频无源互调的研究中,预测无源互调产物的电平是非常重要的。在IM Micro-scope方法的基础上进行了改进,在预测过程中直接引入输入信号经过非线性器件后输出信号的幅度和相位变化模型,给出了详细的数学模型,并实例验证了该方法的正确性。

用IM Microscope方法计算无源互调产物

在微波与射频无源互调的研究中,预测无源互调产物的电平是非常重要的,因为这样可以直观的看出无源互调噪声对于器件或者系统的影响。在众多的预测方法中,IM Microscope 方法具有很多优点。文中在IM Microscope 方法的基础上进行了改进,在预测过程中直接引入输入信号经过非线性器件后输出信号的幅度和相位变化模型,给出了详细的数学模型,这样可以大大减少很多的测量工作。

MicroScope高分辨率成像测井技术及应用

MicroScope随钻高分辨率成像测井技术是斯伦贝谢公司最新推出,能记录超高分辨率电阻率图像UHRI和多个探测深度深钮电阻率成像图,介绍了超高分辨率电阻率成像图UHRI识别岩性及地层结构、识别沉积构造、划分沉积微相、分析古水流与砂体展布等的方法,在南海西部沉积相研究方面发挥了重要作用,是值得推广的新型成像测井技术。

Chalkiness Characters and Scanning Electron Microscope Observation of Rice Grain Endosperm of Japonica Varieties in Southern Henan

Chalkiness characters affect not only the grain appearance,milling,eating and cooking qualities but also the grain nutritional quality in rice,thus it is one of the most important traits in rice. It is very important for us to investigate the relation of the chalkiness formation and the development of endosperm structure and starch granule of different rice varieties. Here,we have investigated the chalkiness characters such as chalkiness rate,chalkiness degree and chalkiness area in 15 japonica rice varieties from southern Henan. Furthermore,the endosperm structure and starch granules of rice grain were also observed with scanning electron microscope. The results showed that the 15 japonica varieties have a significantly linear relationship between the chalkiness rate and chalkiness degree. Among the varieties,the biggest difference is the chalkiness rate,the second is the chalkiness area,and the last is the chalkiness degree. Moreover,there is a certain correlation between the distribution of starch granules,the arrangement of endosperm cells and the occurrence of grain chalkiness in the different rice varieties. For the same variety,the starch granules of chalky and non-chalky grains have obvious difference,while the starch granules from the transparent part of chalky rice and non-chalky rice do not have significant difference. The results would provide useful references for the improvement of grain quality in rice.

Nonlinear control techniques for an atomic force microscope system

Two nonlinear control techniques are proposed for an atomic force microscopesystem. Initially, a learning-based control algorithm is developed for the microcantilever-samplesystem that achieves asymptotic cantilever tip tracking for periodic trajectories. Specifically, thecontrol approach utilizes a learning-based feedforward term to compensate for periodic dynamics andhigh-gain terms to account for non-periodic dynamics. An adaptive control algorithm is thendeveloped to achieve asymptotic cantilever tip tracking for bounded tip trajectories despiteuncertainty throughout the system parameters. Simulation results are provided to illustrate theefficacy and performance of the control strategies.

A novel phase-sensitive scanning near-field optical microscope

Phase is one of the most important parameters of electromagnetic waves. It is the phase distribution that determines the propagation, reflection, refraction, focusing, divergence, and coupling features of light, and further affects the intensity distribution. In recent years, the designs of surface plasmon polariton （SPP） devices have mostly been based on the phase modulation and manipulation. Here we demonstrate a phase sensitive multi-parameter heterodyne scanning near-field opti- cal microscope （SNOM） with an aperture probe in the visible range, with which the near field optical phase and amplitude distributions can be simultaneously obtained. A novel architecture combining a spatial optical path and a fiber optical path is employed for stability and flexibility. Two kinds of typical nano-photonic devices are tested with the system. With the phase-sensitive SNOM, the phase and amplitude distributions of any nano-optical field and localized field generated with any SPP nano-structures and irregular phase modulation surfaces can be investigated. The phase distribution and the interference pattern will help us to gain a better understanding of how light interacts with SPP structures and how SPP waves generate, localize, convert, and propagate on an SPP surface. This will be a significant guidance on SPP nano-structure design and optimization.

Position-resolved Surface Characterization and Nanofabrication Using an Optical Microscope Combined with a Nanopipette/Quartz Tuning Fork Atomic Force Microscope

In this work, we introduce position-resolved surface characterization and nanofabrication using an optical microscope(OM) combined with a nanopipette-based quartz tuning fork atomic force microscope(nanopipette/QTF-AFM) system. This system is used to accurately determine substrate position and nanoscale phenomena under ambient conditions. Solutions consisting of 5 nm Au nanoparticles, nanowires, and polydimethylsiloxane(PDMS) are deposited onto the substrate through the nano/microaperture of a pulled pipette. Nano/microscale patterning is performed using a nanopipette/QTF-AFM, while position is resolved by monitoring the substrate with a custom OM. With this tool, one can perform surface characterization(force spectroscopy/microscopy) using the quartz tuning fork(QTF) sensor. Nanofabrication is achieved by accurately positioning target materials on the surface, and on-demand delivery and patterning of various solutions for molecular architecture.

A stage-scanning laser confocal microscope and protocol for DNA methylation sequencing

Recent understanding of the role of epigenetic regulation in health and disease has necessitated the development of newer and efficient methods to map the methylation pattern of target gene. In this article we report construction of a stage-scanning laser confocal microscope (SLCM) and associated protocol that determines the methylation status of target gene. We have adapted restricted Sanger’s sequencing where fluorescine labeled primers and dideoxy guanine fraction alone are used for target amplification and termination at cytosine positions. Amplified ssDNA bands are separated in 6% denaturing PAGE and scanned using SLCM to sequence the positions of methylated cytosines. We demonstrate that our me- thodology can detect < 100 femtomoles of DNA, and resolve the position of cytosine within ± 2 nucleotide. In a calibration run using a designer DNA of 99 bases, our methodology had resolved all the 11 cytosine positions of the DNA. We have further demonstrated the utility of apparatus by mapping methylation status in the Exon-1 region of a gene, E-Cadherin, in the plasma DNA sample of a healthy subject. We believe our approach constitute a low cost alternative to conventional DNA sequencers and can help develop methylation based DNA biomarkers for the diagnosis of disease and in therapeutics.

Thermal Diffusion of Si Atoms at the Interface of Mo/Si Bilayers Studied with a Soft X-ray Emission Microscope

Thermal diffusion of Si atoms at the interface in Mo/Si multilayers was observed with an imaging type soft X ray emission microscope developed by us. It was possible to observe the diffusion with 0.2nm depth resolution in the direction normal to the interface by comparing the emission intensity for exactly the same position. The diffusion coefficient of Si atoms in Mo at 600℃ was roughly estimated to be 6.0×10 17 cm 2/s.

Confocal Laser Scanning Microscope Evaluation of Early Bacterial Colonization on Zirconium Oxide and Titanium Surfaces:An in vivo Study

To investigate the bacterial colonization on zirconium oxide and titanium surfaces in vivo quantitatively using a confocal laser scanning microscope （CLSM）. Ten samples of zirconium oxide ceramic and commercially pure titanium were fabricated and polished using silicon carbide abrasive paper. One sample from each group was evaluated topographic pattern under a scanning electron microscope. One sample from each group was to evaluate roughness using a profilometer. Eight volunteers were selected. The samples were cemented at the buccal surfaces of upper first molars. All samples were removed after 48 hours, immersed in SYTO-9 and propidium iodide fluorescent to stain for adherent bacteria and obseIved with CLSM. Fewer bacteria were observed in zirconia group than titanium group. However, there was no statistical difference between two groups. The experimental results demonstrate that zirconium oxide may be considered as a promising material for dental implant abutments.

MICRO/NANO-MACHINING ON SILICON SURFACE WITH A MODIFIED ATOMIC FORCE MICROSCOPE

To understand the deformation and removal mechanism of material on nano-scale at ultralow loads,a systemic study on AFM micro/nano-machining on single crystal ailicon is conducted. The results indicate that AFM nano- machining has a precisely dimensional controllability and a good surface quality on nanometer scale.A SEM is adopted to observe nano-machined region and chips,the results indicate that the material removal mechanisms change with the applied normal load. An XPS is used to analyze the changes of chemical composition inside and outside the nano-machined region respectively.The nano-indentation which is conducted with the same AFM diamond tip on the machined region shows a big discrepancy compared with that on the macro-scale. The calculated results show higher nano-hardness and elastic modulus than normal values .This phenomenon on be regarded as the indentation size effect(ISE).

Observation of MWCNTs with low-energy electron point source microscope

The low-energy electron point source （LEEPS） microscope, which creates enlarged projection images with lowenergy field emission electron beams, can be used to observe the projection image of nano-scale samples and to characterize the coherence of the field emission beam. In this paper we report the design and test operation performance of a home-made LEEPS microscope. Multi-walled carbon nanotubes （MWCNTs） synthesized by the CVD method were observed by LEEPS microscope using a conventional tungsten tip, and projection images with the magnification of up to 10^4 was obtained. The resolution of the acquired images is -10 nm. A higher resolution and a larger magnification can be expected when the AC magnetic field inside the equipment is shielded and the vibration of the instrument reduced.

Surface Electromechanical Coupling on DLC Film with Conductive Atomic Force Microscope

Diamond-like carbon (DLC) film composed of microscopically insulation but microscopically a mixture of conducting (sp2) and insulating (sp3) phases was discussed on the local modification with a conductive atomic force microscope (C-APM). Especially, a topographic change was observed when a direct current (DC) bias-voltage was applied to the DLC film. Experimental results show that a nanoscale pit on DLC surface was formed when applying a positive 25 V on DLC film. According to the interacting force between CoCr-coated microelectronic scanning probe (MESP) tip and DLC surface, as well as the Sondheimer oscillation theory, the 'scalewing effect' of the pit was explained. Electromechanical coupling on DLC film suggested that the depth of pits increased with an increase of load applied to surface when the cantilever-deflected signal was less than a certain threshold voltage.

Concanavalin A-induced changes in lysosomal morphology of macrophages under confocal microscope

Changes in lysosomal morphology of cultured mouse peritoneal macrophages after stimu1ation by Concanavalin A (Con A) were observed with a laser scanning confocal microscope (LSCM). A series of images were obtained including phase-contrast images, optical sectioning images, 3-dimensional reconstruction images. The changes of lysosomal fluorescence intensity and pH were measured. It was found that macrophage lysosomes were Cllstributed mainly at the periphery of the cells in resting conditions, the lysosomal area containing fluorescence probe became markedly enlarged after stimulation by Con A for 30 min, and the fluorescence intensity in the medium increased about 15 min after suggesting that Con A could induce outflow of the fluorescence probe within the macrophage lysosomes. The lysosomal pH rose from 4. 6 to 5. 7 in 7 min after Con A was added, and maintained at that level hereafter.

Research on a Diamond Tip Wear Mechanism in Atomic Force Microscope-based Micro/nano-machining

The object is to investigate the wear of an atomic force microscope (AFM) diamond tip when conducting micro/nano machining on single crystal silicon surface. The experimental research and theoretical analysis were carried out on the worn tip in terms of wear rate, wear mechanism and the effect of the tip wear on micro machining process. The wear rate was calculated as 1.7(10～10mm 3/(N·m) by using a theoretical model combined with the experimental results. Through an integration of an AFM observation on the worn tip features with the FEM simulation of the stress distribution, in addition to the unit cutting force calculation on the AFM diamond tip, the wear mechanism of the AFM diamond tip was concluded as mainly chemical wear, and the wear process was also elaborated as well.

OBSERVATION OF THE EFFECT OF S－（－）USNIC ACID SODIUM ON TRICHOMONAS VAGINALIS BY TRANSMISSION ELECTRON MICROSCOPE

This paper reports the ultrastructural changes of trichomonas vaginaiis (T. vag. ) under the action of s-(-)usnic acid sodium in vitro. These changes can be shown by the following results:At first, the degranulation of the rough endoplasmic reticulum (RER) took place, the polyribosomes disaggregated. And then, the Golgi complex and the endoplasmic reticulum dilated. The cytoplasmic matrix presented and inhomogeneous apperance. Finally, the biomembrane loosed and fractured.The cell nuclei presented the karyorrhexis.

At wavelength coherent scatterometry microscope using high-order harmonics for EUV mask inspection

In this review,we describe our research on the development of the 13.5 nm coherent microscope using high-order harmonics for the mask inspection of extreme ultraviolet(EUV)lithography.EUV lithography is a game-changing piece of technology for high-volume manufacturing of commercial semiconductors.Many top manufacturers apply EUV technology for fabricating the most critical layers of 7 nm chips.Fabrication and inspection of defect-free masks,however,still remain critical issues in EUV technology.Thus,in our pursuit for a resolution,we have developed the coherent EUV scatterometry microscope(CSM)system with a synchrotron radiation(SR)source to establish the actinic metrology,along with inspection algorithms.The intensity and phase images of patterned EUV masks were reconstructed from diffraction patterns using ptychography algorithms.To expedite the practical application of the CSM,we have also developed a standalone CSM,based on high-order harmonic generation,as an alternative to the SR-CSM.Since the application of a coherent 13.5 nm harmonic enabled the production of a high contrast diffraction pattern,diffraction patterns of sub-100 ns size defects in a 2D periodic pattern mask could be observed.Reconstruction of intensity and phase images from diffraction patterns were also performed for a periodic line-and-space structure,an aperiodic angle edge structure,as well as a cross pattern in an EUV mask.