Download Cell Imaging Techniques: Methods and Protocols by Diana P. Bratu (auth.), Douglas J. Taatjes, Brooke T. PDF

By Diana P. Bratu (auth.), Douglas J. Taatjes, Brooke T. Mossman (eds.)

A various choice of cutting-edge equipment for the microscopic imaging of cells and molecules. The authors disguise a large spectrum of complimentary suggestions, together with such tools as fluorescence microscopy, electron microscopy, atomic strength microscopy, and laser scanning cytometry. extra effectively reproducible protocols on confocal scanning laser microscopy, quantitative computer-assisted snapshot research, laser-capture microdissection, microarray photograph scanning, near-field scanning optical microscopy, and mirrored image distinction microscopy around out this eclectic number of state-of-the-art imaging concepts now to be had. The authors additionally speak about preparative tools for debris and cells via transmission electron microscopy.

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Extra resources for Cell Imaging Techniques: Methods and Protocols

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Now that multiphoton microscopes are in wide use, it seems inescapable that the associated technique of SHG microscopy will grow steadily more important in future years. 4. Notes 1. The process of second harmonic generation (SHG) has been summarized in a recent paper by Gauderon et al. (2). Briefly, as electromagnetic radiation propagates through matter, the electric field (E) exerts forces on the sample’s internal charge distribution. The consequent redistribution of charge generates an additional field component.

Fluorescence always involves some loss of energy in the sample, and the fact that electrons are raised to excited states means that bleaching is inevitable. Harmonic Imaging of Collagen 17 Table 1 Comparison of SHG and TPF SHG Exactly double the original frequency Largely frequency independent Virtually instantaneous, approx 1 fs Propagated forward Coherent (in phase with exciting light) No energy loss or damage Requires short laser pulses TPF Spectrum of frequencies less than two times the original Strongly frequency dependent Lifetime in nanoseconds Propagated in all directions Incoherent Always energy loss and associated damage Requires short laser pulses SHG dissipates no energy in the sample and there is no excitation or bleaching.

A) Forward-propagated signal, almost exclusively SHG at 410 nm, although detected over a wide spectral range (400–550 nm). (B) Back-propagated signal in the spectral range 500- to 550-nm showing aldehyde-induced autofluorescence. The photomultiplier tube voltage was set 100 V higher than in (A) in order to pick up the much weaker autofluorescence. Previous collection of an image at a higher zoom setting has caused slight bleaching in the fluorescent image (arrowed), but the second-harmonic image is unaffected.

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