Image Formation in the Scanning Microscope

Sheppard, Colin J. R.; Choudhury, Arup

doi:10.1080/713819421

Cited by 451 publications

(206 citation statements)

References 9 publications

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“…A beam splitter and second photomultiplier permits dualwavelength imaging of specimens stained with fluorescein and Texas red. This technique is called confocal because the image of the illumination pinhole and the detector pinhole have a common focus in the specimen (23). The digitized photomultiplier output at each point in the raster scan is assembled into an image, up to 768 x 512 pixels, by means of a frame store in the microcomputer.…”

Section: Methodsmentioning

confidence: 99%

Three-Dimensional Imaging of Intact Isolated Islets of Langerhans With Confocal Microscopy

Brelje

Sorenson

1989

Diabetes

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We present a new technique for analyzing the three-dimensional structure of intact isolated islets of Langerhans. Adult rat and human islets were stained with whole-mount immunofluorescence techniques and optically sectioned with a confocal microscope. This has several advantages over traditional methods: 1) the technical difficulties in serial sectioning and handling the large numbers of sections are avoided, 2) optical sectioning by confocal microscopy gives improved resolution and strongly suppresses light from out-of-focus structures, and 3) entire islets can be rapidly imaged for the presence of positive staining. This new technique should facilitate the study of the three-dimensional structure of islets of Langerhans. Diabetes 38:808-14,1989 T he three-dimensional structure of islets of Langerhans is important in understanding the regulatory mechanisms involved in controlling the different islet cell types. Since the first immunofluorescent localization of insulin to p-cells by Lacy and Davies (1,2), glucagon to the a-cell by Baum et al. (3), somatostatin to the 5-cell by several investigators (4-7), and pancreatic polypeptide (PP) to a distinct cell type by Larsson and coworkers (8-10), fluorescence microscopy has been an important technique in investigating the structure of islets.However, a severe limitation with conventional fluorescence microscopy has been fluorescent background resulting from out-of-focus structures, which reduces image contrast and therefore limits the amount of information that can be obtained from in-focus structures. To overcome this limitation, it has been necessary to examine thin sections of

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Section: Methodsmentioning

confidence: 99%

Three-Dimensional Imaging of Intact Isolated Islets of Langerhans With Confocal Microscopy

Brelje

Sorenson

1989

Diabetes

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“…Early presentation of the imaging abilities and the initial improved confocal was displayed in publication by Brakenhoff (1979). Sheppard et al (1977) and Brakenhoff (1979) both employed laser as a light source. They also used a mechanical scanning that moves the specimens through the stationary confocal point.…”

Section: Cslmmentioning

confidence: 99%

“…According to Brakenhoff et al, (1988) Confocal scanning light microscopy forms a bridge between conventional light microscopy with its limited resolution (but capable of imaging hydrated, possibly live, specimens) on the one side, and electron microscopy with its higher resolution on the other. The principal of the basic confocal microscope has been explained by Sheppard and Choudhury (1977). This microscope might be operated in two techniques: reflection or in fluorescence.…”

Section: Cslmmentioning

confidence: 99%

Microwave processing of meat

Yarmand¹,

Rad²

2011

Microwave Heating

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“…It has been demonstrated that the theoretically expected improved imaging (Sheppard & Choudhury, 1977) in confocal scanning electron microscopy (CSLM) can indeed be realized for the high aperture immersion optics, required for high resolution light microscopy (Brakenhoff et al, 1979). The increase in resolution for point objects is by a factor of 1.4 as compared with conventional microscopy.…”

Section: Introductionmentioning

confidence: 99%

Imaging modes in confocal scanning light microscopy (CSLM)

Brakenhoff

1979

Journal of Microscopy

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SUMMARYThe optical arrangement for confocal scanning light microscopy can be incorporated in various imaging modes. Light microscopical specimens can be imaged with contrast enhanced, under y-control, inverted, etc. In interference, conditions can be set such that either pure phase or pure amplitude images result. Stereoscopic images at arbitrary aspect ratios can be realized in CSLM by electronic processing of the data obtained when the specimen is sampled with more than one confocal point concurrently. Also forms of differential imaging either amplitude or phase are possible. The coupling of these imaging modes with the improved resolving powers of CSLM results in some unique imaging opportunities, especially of value for high resolution light microscopy of living specimens.

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Image Formation in the Scanning Microscope

Cited by 451 publications

References 9 publications

Three-Dimensional Imaging of Intact Isolated Islets of Langerhans With Confocal Microscopy

Three-Dimensional Imaging of Intact Isolated Islets of Langerhans With Confocal Microscopy

Microwave processing of meat

Imaging modes in confocal scanning light microscopy (CSLM)

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