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PubMed2003

Enhancement of lateral resolution in confocal self-interference microscopy.

We describe confocal self-interference microscopy with enhanced lateral resolution. A uniaxial anisotropic crystal is used to cause interference between two linearly polarized beams that are reflected from the same pointlike object in the focal plane of the objective lens. Theory and the optimal design that maximizes the sensitivity of the interference signal are presented. A numerical experiment shows a 38% decrease in the lateral FWHM for simple confocal self-interference microscopy.

Journal Article↗
PubMed1976

Degranulation of chicken heterophil leucocytes during phagocytosis, studied by phase contrast and interference microscopy.

The dynamic aspects of degranulation of chicken heterophil leucocytes during phagocytosis have been studied by phase contrast and interference microscopy. Both standard (8 fps) and high-speed (400 fps) cine-photomicrographic recordings of this process under phase contrast are presented. Lysis of individual granules is usually completed in less than 60 milliseconds. During lysis of each granule a rounded phase dense body is ejected into the phagocytic vacuole. Measurements made by interference microscopy show that there is usually a substantial fall from a protein concentration of c. 100 per cent. w/v for intact granules to a concentration of 16 per cent. w/v for the vacuole resulting from their lysis; this can only be explained by a rapid intake of water into the granule matrix following membrane fusion. The intake of water that accompanies granule lysis causes swelling of the granule matrix, and is thought to explain the mechanism of ejection of the phase dense body. Granule lysis is not dependent on the intake of water, since occasional vacuoles have been observed which showed no fall in protein concentration relative to the intact granules. The membrane around the intact granule effectively excludes the entry of water into the concentrated hygroscopic granule matrix, but once membrane fusion occurs this barrier to the entry of water is usually lost.

Animals↗
PubMed1993

Phase-shifting interference microscopy applied to the analysis of cell behaviour.

The theory of phase-shifting interferometry is not new but it is only recently, with the advent of solid-state detector arrays and fast image processors, that it has become a practical imaging technique. In conjunction with transmission interference microscopy, phase-shifting presents a new way of introducing contrast into the images of transparent microscopic objects such as cultured cells. An earlier paper from our laboratory has emphasised the advantages of transmission interference microscopy over phase contrast or differential interference contrast microscopy for the computerised analysis of cell behaviour. Phase-shifting greatly improves the accuracy, long-term stability and range of application of this technique but it has not previously, to our knowledge, been combined with transmission interference microscopy for the study of cultured cells. The resulting image is especially well suited to quantitative analysis by computer since it is a direct representation of the distribution of non-aqueous cellular material in the specimen. The image is not degraded by uneven illumination; by heterogeneous sensitivity of the detector array; or by differential absorption of light in the optics or specimen. Our main purpose in developing the method is to obtain sequences of images of the motile behaviour of cells in culture for analysis by computer. This type of analysis is potentially a powerful tool for studying the motile responses of cells and the operation and control of their locomotory machinery. Not only can the method be used for studying cell translocation and the dynamics of intracellular movement of non-aqueous material, but it is now possible to study in detail the time course of growth in individual cultured cells.

Animals↗