Single-shot phase contrast microscopy using polarisation-resolved differential phase contrast

We present a robust, low-cost single-shot implementation of differential phase microscopy utilising a polarisation-sensitive camera to simultaneously acquire four images from which phase contrast images can be calculated. This polarisation-resolved differential phase contrast (pDPC) microscopy technique can be easily integrated with fluorescence microscopy.     

Morphological study of the bovine blastocyst with phase contrast microscopy

The morpholoqical appearance of 40 bovine blastocysts. collected from single ovulating heifers 7 days after oestrus was evaluated with the aid of a phase contrast inverted microscope. Out of 40 blastocysts, 17 were classified normal (N), 14 in the process of degeneration (IPD) and 9 degenerated (D). There was no difference in the mean diameters of the three groups of blastocysts, whereas the mean diameters of cell mass was bigger in N than IPD and D blastocysts. This difference was highly significant (P < .005). The mean number of cells was greater in normal than in abnormal (99 vs 58). This difference was highly significant (P < .005). The results reported here indicate that the criteria used for classification of the morphological appearance of the 7 day old bovine blastocyst reflected the degree of cellular organization and the number of cells.     

Zernike phase contrast electron microscopy of ice-embedded influenza A virus

The ultrastructure of the frozen-hydrated influenza A virus was examined by Zernike phase contrast electron microscopy. Using this new microscopy, not only lipid bilayers but also individual glycoprotein spikes on viral envelopes were clearly resolved with high contrast in micrographs taken in focus. In addition to spherical and elongated virions, three other classes of virions were distinguished on the basis of the features of their viral envelope: virions with a complete matrix layer, which were the most predominant, virions with a partial matrix layer, and virions with no matrix layer under the lipid bilayer. About 450 glycoprotein spikes were present in an average-sized spherical virion. Eight ribonucleoprotein complexes, that is, a central one surrounded by seven others, were distinguished in one viral particle. Thus, Zernike phase contrast electron microscopy is a powerful tool for resolving the ultrastructure of viruses, because it enables high-contrast images of ice-embedded particles free of contrast transfer function artifacts that can be a problem in conventional cryo-electron microscopy.     

Observations on the cytoplasmic membranes of testicular cells,examined by phase contrast and electron microscopy

In freshly isolated cells of the guinea pig germinal epithelium examined with phase contrast, dark contours are seen in the cytoplasm that appear to be optical sections of the cisternae of the endoplasmic reticulum. These increase in contrast, in number, and in linear extent with increasing time up to 4 hours after isolation of the cells from the testis. During this period, cisternae originally present in the cells are extended and new ones appear to be formed by coalescence of tubular and vesicular elements of the reticulum. The cisternae become associated in parallel array and ultimately form elaborate concentric systems resembling structures that have often been interpreted as intracellular "myelin figures." Until now our knowledge of the endoplasmic reticulum has been based largely upon electron micrographs. The observation that the cisternae are visible in certain cell types under phase contrast optics opens the way for experimental investigations on the behavior of this class of cytoplasmic membranes in living cells.     

Advantages of intermediate X-ray energies in Zernike phase contrast X-ray microscopy

Understanding the hierarchical organizations of molecules and organelles within the interior of large eukaryotic cells is a challenge of fundamental interest in cell biology. Light microscopy is a powerful tool for observations of the dynamics of live cells, its resolution attainable is limited and insufficient. While electron microscopy can produce images with astonishing resolution and clarity of ultra-thin (< 1 μm thick) sections of biological specimens, many questions involve the three-dimensional organization of a cell or the interconnectivity of cells. X-ray microscopy offers superior imaging resolution compared to light microscopy, and unique capability of nondestructive three-dimensional imaging of hydrated unstained biological cells, complementary to existing light and electron microscopy.     

Newly designed,simple relief phase contrast for microscopy of microorganisms

A new method providing a relief phase contrast for investigation of microorganisms by optical microscopy used a neutral filter Zeiss NG 10/1 that could be controllably slid at a certain azimuthal angle below the aperture condenser diaphragm of the microscope phase contrast. Two ways of application are described depending on the type of the microscope: (1) in a special holder, and (2) fixed on a rubber ring. The device enabled us to obtain excellent results in the area of both optical microscopy and microphotography. With the microorganisms visualized, a better resolution, higher contrast and a significant 3D effect were obtained; outer morphology and organelles (chloroplasts, nuclei, granules, oil reserve vacuoles, etc.) could also be investigated.