Method for the detection of dyes in fluorescence microscopy
AssigneeCarl Zeiss Jena GmbH
InventorsRalf Wolleschensky, Gunter Moehler
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Abstract
A method for optical detection of characteristic quantities of the wavelength-dependent behavior of an illuminated specimen, such as the emission behavior and/or absorption behavior, preferably the fluorescence and/or enzyme fluorescence comprises determining at least one spectral centroid and/or a maximum of the emission radiation and/or of the absorbed radiation. An arrangement for performing the method is disclosed.
Claims84
A method for optical detection of quantities of different dyes in a specimen by detecting the wavelength-dependent behavior of an illuminated specimen in an image generating arrangement using spectra which contain overlapping dye spectra from different dyes which are simultaneously examined, the wavelength-dependent behavior including the emission behavior, absorption behavior, fluorescence, luminescence, phosphorescence, enzyme-active light emission, naturally occurring dye fluorescence from fluorescent proteins or enzyme-active fluorescence of the illuminated specimen, comprising:
- determining at least one spectral centroid position for different dye spectra detected by a spectral detector wherein a characteristic dye spectra is detected for at least one image point of the specimen;
- and
- applying a characteristic spectral weighting function to the characteristic dye spectra that is detected to separate overlapping dye components and to calculate the intensity of each dye component per desired image point.
An arrangement for optical detection of characteristic quantities of the wavelength-dependent behavior of an illuminated specimen using spectra which contain overlapping dye spectra from different dyes which are simultaneously examined, the wavelength-dependent behavior including the emission behavior, absorption behavior, fluorescence, luminescence, phosphorescence, enzyme-active light emission, naturally occurring dye fluorescence from fluorescent proteins or enzyme-active fluorescence of an illuminated specimen, comprising:
- an illumination source that provides an illumination for a specimen;
- a spectral detector that receives detection radiation coming from the illuminated specimen;
- a computer for determining at least one spectral centroid position for different dye spectra detected by the spectral detector wherein a characteristic dye spectra is detected for at least one image point of the specimen and the computer applies a characteristic spectral weighting function to the characteristic dye spectra that is detected to separate overlapping dye components and to calculate the intensity of each dye component per desired image point.
Dependencies are inferred from the claim text.