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CERTAIN TECHNICAL ASPECTS OF FLUORESCENCE MICROSCOPY AND THE COONS FLUORESCENT ANTIBODY TECHNIQUE

Authors: J E, TOBIE;

CERTAIN TECHNICAL ASPECTS OF FLUORESCENCE MICROSCOPY AND THE COONS FLUORESCENT ANTIBODY TECHNIQUE

Abstract

A fluorescence microscope possessing an efficient ultraviolet light source, the 1000 watt A-H6, is described with special reference to its application in studies concerned with the fluorescent antibody technique and the tissue localization of the tetracyclines. The spectral characteristics of the light source are discussed and methods outlined whereby combinations of certain ultraviolet transmitting and absorbing filters can be used for maximum stimulation and visualization of fluorescent substances in tissue specimens. In fluorescent antibody studies when fluorescein isocyanate is used as the fluor, superior results have been obtained by excitation of tissue sections with blue-violet radiation at a wavelength band of 390-440 mµ rather than with ultraviolet radiation at 360-370 mµ. The two outstanding advantages of the longer wavelength stimulation are: 1) The increased fluorescence and consequent visual enhancement; and 2) the greater cellular detail for visual orientation and photographic clarity thus obtained. In contrast to this, the tetracyclines were more effectively stimulated in sections of bone with ultraviolet radiation at wavelengths in the 360-370 mµ band. A method of quick-freezing fresh tissues in petroleum ether at approximately –65° C has been found more satisfactory for preserving the cellular architecture of tissues used in the fluorescent antibody technique. The operation of a rotary microtome in a cryostat at sub-freezing temperatures has been improved by special lubrication with the result that sections can be cut at a microtome setting of 3 µ. With these thin sections, it is possible to observe more minute cytological details and to localize fluorescent substances more accurately.

Related Organizations
Keywords

Microscopy, Microscopy, Fluorescence, Fluorescent Antibody Technique, Humans, Antibodies, Fluorescence

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selected citations
These citations are derived from selected sources.
This is an alternative to the "Influence" indicator, which also reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Citations provided by BIP!
popularity
This indicator reflects the "current" impact/attention (the "hype") of an article in the research community at large, based on the underlying citation network.
BIP!Popularity provided by BIP!
influence
This indicator reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Influence provided by BIP!
impulse
This indicator reflects the initial momentum of an article directly after its publication, based on the underlying citation network.
BIP!Impulse provided by BIP!
42
Average
Top 1%
Top 10%
bronze