Edge detection,three-dimensional cell boundary reconstruction and volume and surface area estimation from differential interference contrast images

Three-dimensional (3-D) cell morphology is important for the understanding of cell function and can by quantified in terms of volume and surface area. Differential interference contrast (DIC, or Nomarski) imaging can enable cell edges to be clearly visualized in unstained tissue due to the slight difference in refractive index between aqueous media and cytoplasm. DIC is affected in only one direction - the direction of the optical shear. A 1-D edge detector was used in that direction with a scale length equal to that of an in-focus edge to highlight cell boundaries. By comparison with the signal from the edge detector on an out-of-focus slice, the in-focus slices could be segmented and, after noise suppression, cell outlines obtained. A voxel paradigm was used to calculate cell volume and differential geometry was used for surface area estimation. We applied this approach to obtain 3-D dimensional information by optical sectioning of motile Amoeba proteus.     

基于MATLAB的Nomarski棱镜优化设计

根据落射式微分干涉相衬显微镜中Nomarski棱镜的光学原理,通过几何光学运算,推导出棱镜主要结构参数的理论关系式,利用MATLAB程序将关系式转化为二维曲线,再根据关系曲线的变化趋势来确定适合于Nomarski棱镜主要结构的参数取值范围,从而实现优化设计的目的。     

A simple method allowing DIC imaging in conjunction with confocal microscopy

Current optical methods to collect Nomarski differential interference contrast (DIC) or phase images with a transmitted light detector (TLD) in conjunction with confocal laser scanning microscopy (CLSM) can be technically challenging and inefficient. We describe for the first time a simple method that combines the use of the commercial product QPm (Iatia, Melbourne Australia) with brightfield images collected with the TLD of a CLSM, generating DIC, phase, Zernike phase, dark-field or Hoffman modulation contrast images. The brightfield images may be collected at the same time as the confocal images. This method also allows the calculation of contrast-enhanced images from archival data. The technique described here allows for the creation of contrast-enhanced images such as DIC or phase, without compromising the intensity or quality of confocal images collected simultaneously. Provided the confocal microscope is equipped with a motorized z-drive and a TLD, no hardware or optical modifications are required. The contrast-enhanced images are calculated with software using the quantitative phase-amplitude microscopy technique ( Barone-Nugent et al., 2002 ). This technique, being far simpler during image collection, allows the microscopist to concentrate on their confocal imaging and experimental procedures. Unlike conventional DIC, this technique may be used to calculate DIC images when cells are imaged through plastic, and without the use of expensive strain-free objective lenses.     

Confocal differential interference contrast (DIC) microscopy: including a theoretical analysis of conventional and confocal DIC imaging

A technique for obtaining differential interference contrast (DIC) imaging using a confocal microscope system is examined and its features compared to those of existing confocal differential phase contrast (DPC) techniques as well as to conventional Nomarski DIC. A theoretical treatment of DIC imaging is presented, which takes into account the vignetting effect caused by the finite size of the lens pupils. This facilitates the making of quantitative measurements in DIC and allows the user to identify and select the most appropriate system parameters, such as the bias retardation and lateral shear of the Wollaston prism.     

光学玻璃材料的破坏和折射率变化的测量研究

介绍了一套用Nomarski干涉仪测量、面阵CCD摄像、微机实时监控记录光学玻璃材料在激光作用下破坏的装置,并对试验室原有的Nomarski干涉仪进行了优化设计,提高了它的探测灵敏度,使整套设备更加小型化.     

Nomarski偏振光干涉仪与短脉冲相干光源

通过详细计算给出用超短光脉冲作相干光源的偏振光干涉仪中Ｗｏｌｌａｓｔｏｎ棱镜的设计参数，并仔细分析了棱镜在光路中处于不同位置时对干涉仪性能的影响。     

Measurement-based evaluation of optical pathlength distributions reconstructed from simulated differential interference contrast images

Recently a method was presented for reconstructing optical pathlength distributions (OPDs) from images of weak phase objects obtained by a conventional differential interference contrast (DIC) microscope. A potential application of this technique is the determination of the mass of biological objects: by integrating the optical pathlength over the projected surface of the image of an object, a measure of the dry mass, i.e. the total mass of all solid constituents present in the object, is obtained. To assess the possibilities of DIC microscopy for this application, simulations were performed on computer-generated DIC images of objects of various sizes, shapes and orientation angles. After reconstructing the OPDs from these images, the integrated optical pathlength of each of the test objects was determined, and compared with the expected results. The parameter settings used in the reconstruction algorithm were found to be very important in obtaining a reliable measurement. Using optimal parameter settings, errors in the integrated OPD could be limited to a few per cent for circular objects within the investigated size range. For non-circular objects, however, the orientation angle of the object relative to the lateral shift was found to influence the measured values. Ellipses with their long axes perpendicular to the shift direction had a significantly higher integrated OPD than ellipses orientated parallel to the shift. By adjusting the reconstruction parameters the effect could be limited, but complete elimination of the artefact was not possible within the parameter range investigated.     

用Nomarski干涉仪与数字相机研究光学材料的破坏

介绍自行研制的一台新型光学材料破坏的测量装置.它由数字相机与Nomarski偏光干涉仪组成.并与计算机接口.从而实现光学材料破坏测量过程中的数字化.并用它对光学材料破坏进行初步测量和分析.实验结果与分析相符.     

掺碳GaAs生长特性的研究

采用以碳纤维为碳源的固态源MBE技术，生长了不同厚度的重接碳GaAs以及具有不同表层厚度的δ碳掺杂GaAs，通过Nomarski干涉显微镜和原子力显微镜（AFM）对样品表面形貌的观察，分析了挨碳GaAs的生长过程和各种缺陷的产生，提出碳的掺入导致了GaAs材料的三维岛状生长，促进了各种缺陷的力生。提出了通过改善生长条件减少缺陷的途径。     

Astronomical hypersensitization techniques applied to photomicrography: Kodak Technical Pan Film 2415

Kodak Technical Pan Film 2415 is a valuable recording medium because of its unusually fine grain and high resolution. Hypersensitization techniques developed by astronomers can increase its normally low speed several-fold in the range of exposures commonly employed in microscopy. The speed gains are even larger for very long exposures such as those encountered in fluorescence work. Troublesome effects from reciprocity failure are virtually eliminated. When active development is added to hypersensitization, well-exposed negatives can be obtained at ISO indexes as high as 1600. As in astronomy, the same general techniques can be applied to a wide variety of emulsions.