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Working with the confocal scanning UV‐laser microscope: specific DNA localization at high sensitivity and multiple‐parameter fluorescence
Author(s) -
Montag Markus,
Kukulies Jörg,
Jörgens Reinhard,
Gundlach Heinz,
Trendelenburg Michael F.,
Spring Herbert
Publication year - 1991
Publication title -
journal of microscopy
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.569
H-Index - 111
eISSN - 1365-2818
pISSN - 0022-2720
DOI - 10.1111/j.1365-2818.1991.tb03172.x
Subject(s) - confocal , dapi , fluorescence , microscope , confocal microscopy , fluorescence microscope , laser , microscopy , laser scanning , optics , chromatin , ultraviolet , biophysics , microscope slide , materials science , staining , dna , biology , physics , genetics
SUMMARY The use of fast‐staining DNA‐specific dyes such as DAPI or Hoechst 33342/33258 has been a major problem for confocal scanning laser microscopy (CSLM) studies of intranuclear chromatin organization. Moreover, the availability of a confocal ultraviolet scanning laser microscope configuration, which allows an excitation at wavelengths of 364 nm as well as 488, 514 and 543 nm, is a prerequisite for single as well as multiple fluorescence parameter studies, especially if these studies are concerned with the precise localization of intranuclear signals. Here we report the characteristics and application of a CSLM, which was adapted for UV‐excitation and therefore enables comparison of the spatial distribution of several types of signals within one preparation. In addition to multiple‐parameter studies, we have also investigated the sensitivity of the system with regard to the identification of the double‐stranded DNA of lampbrush chromosome loops in germinal vesicles of amphibian oocytes.