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Ultrathin fluorescent layers for monitoring the axial resolution in confocal and two‐photon fluorescence microscopy
Author(s) -
Martin Schrader,
Ulrich G. Hofmann,
Stefan W. Hell
Publication year - 1998
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 - 10.1046/j.1365-2818.1998.00361.x
Subject(s) - fluorescence , microscope , confocal , fluorescence microscope , two photon excitation microscopy , microscopy , optics , resolution (logic) , materials science , ammonium bromide , optical microscope , confocal microscopy , fluorescence lifetime imaging microscopy , chemistry , analytical chemistry (journal) , scanning electron microscope , physics , chromatography , pulmonary surfactant , biochemistry , artificial intelligence , computer science
Monomolecular films of polymerized dimethyl‐bis[pentacosadiinoic‐oxyethyl] ammonium bromide (EDIPAB) provide one‐ and two‐photon excited fluorescence that is sufficiently high to quantify the axial resolution of 3‐D fluorescence microscopes. When scanned along the optical axis, the fluorescence of these layers is bright enough to allow online observation of the axial response of these microscopes, thus facilitating alignment and fluorescence throughput control. The layers can be used for directly measuring and monitoring the axial response of 4Pi‐confocal microscopes, as well as for their initial alignment and phase adjustment. The proposed technique has the potential to supersede the conventional technique of calculating the derivative of the axial edges of a thick fluorescent layer. Coverslips with EDIPAB‐layers can be used as substrates for the cultivation of cells.