High dynamic range scanning with reduced channel cross-talk

A fluorescence detection system, comprising:

• a first illumination source that emits at a first illumination frequency;
• a second illumination source that emits at a second illumination frequency;
• a first detector element configured to detect radiation at a first fluorescence frequency range;
• a second detector element configured to detect radiation at a second fluorescence frequency range;
• a target platform having a sample with two fluorescent species, wherein a first species absorbs illumination at the first illumination frequency and emits at the first fluorescence frequency, and wherein a second species absorbs illumination at the second illumination frequency and emits at the second fluorescence frequency;
• optical elements configured to combine and focus illumination of the first and second sources onto the sample;
• optical elements configured to direct and focus fluorescence light emitted by the sample onto the first and second detector elements;
• and
• an intelligence module adapted to receive and process signals received from the first and second detectors,
• wherein, during operation, at least one pixel location on the sample platform is simultaneously illuminated by the first and second sources in a first mode and in a second mode, and wherein fluorescence emitted by the first and second species is simultaneously detected by the first and second detectors in the first mode and in the second mode,
• wherein in the first mode, the first illumination source is set to a high power setting and the first detector is set to a high gain setting, and the second illumination source is set to a low power setting and the second detector is set to a low gain setting, and
• wherein in the second mode, the first illumination source is set to a low power setting and the first detector is set to a low gain setting, and the second illumination source is set to a high power setting and the second detector is set to a high gain setting,
• wherein in the first mode, each detector outputs a first intensity value, and wherein in the second mode, each detector outputs a second intensity value, and
• wherein the processor, for each detector, processes the first and second intensity values to produce a merged image value for the at least one pixel location.

A method of reducing or eliminating cross talk in a fluorescence detection system having two sources that emit illumination at different wavelengths, a sample platform having at least two fluorescent species and two detection channels, the method comprising:

• a) setting the illumination power of a first source to a high power level and the illumination power of a second source to a low power level;
• b) setting the gain of a first detector to a high gain level and the gain of a second detector to a low gain level;
• c) simultaneously illuminating at least one of a plurality of pixel locations of the sample platform with illumination from the first and second sources, wherein a first fluorescent species absorbs light from the first source and emits at a first fluorescence wavelength, and wherein a second fluorescent species absorbs light from the second source and emits at a second fluorescence wavelength;
• and
• d) simultaneously detecting a first intensity value for each detection channel, wherein a first detector detects the first fluorescence wavelength range and a second detector detects the second fluorescence wavelength range;
• and thereafter
• e) setting the illumination power of the first source to a low power level and the illumination power of the second source to a high power level;
• f) setting the gain of the first detector to a low gain level and the gain of the second detector to a high gain level;
• g) simultaneously illuminating the at least one pixel location with illumination from the first and second sources;
• h) simultaneously detecting a second intensity value for each channel;
• and thereafter
• i) for each channel, processing the first and second intensity values to produce a merged image value for the at least one pixel location.