Patent ID: 8294897

Claim:
A method for imaging a sample using a microscope, the method comprising: an illumination step in which a sample is illuminated with excitation light via an illuminating beam path; and a recording step in which light emitted from the sample is recorded via a detection beam path; wherein at least one adjustable beam splitter having an adjustable threshold wavelength between reflection and transmission is arranged in the detection beam path or/and in the illuminating beam path; wherein light emitted from the sample is detected in at least one detection channel; wherein, if the adjustable beam splitter is arranged in the detection beam path, the emitted light is split by the beam splitter and either emitted light passing through the beam splitter or light reflected by the beam splitter enters the detection channel; wherein, if the adjustable beam splitter is arranged in the illuminating beam path, a light spectrum of the light source is split by the beam splitter for varying the spectrum of the excitation; wherein the adjustable beam splitter is one of a low-pass filter and a high-pass filter; wherein, for at least one predetermined sample region, a signal intensity of light detected in the at least one detection channel is recorded for a plurality of threshold wavelengths set at the adjustable beam splitter to obtain a signal/threshold-dependency of the predetermined sample region; and wherein, from the signal/threshold-dependency or dependencies, at least one setting wavelength is determined, the setting wavelength representing a setup of the adjustable beam splitter for resolving fluorochromes with overlapping emission spectra, and the setting wavelength being a wavelength where emission lines of the fluorochromes show comparable intensities.