Fluorescence excitation and detection system and method

Abstract

System including an excitation assembly that is configured to excite a sample with first and second wavelengths during first and second excitation events, respectively, of an illumination/detection cycle. The sample provides emission light that includes different first, second, third, and fourth spectral patterns in response to the first and second wavelengths. The system also includes a first detection camera that detects the first spectral pattern during a first measurement phase of the illumination/detection cycle. The first detection camera detects the third spectral pattern during a second measurement phase of the illumination/detection cycle. The system also includes a second detection camera that detects the second spectral pattern during the first measurement phase. The second detection camera detects the fourth spectral pattern during the second measurement phase.

1 . (canceled) 
     
     
         2 . A system comprising:
 an excitation assembly configured to excite a sample with first and second wavelengths during first and second excitation events, respectively, of an illumination/detection cycle, wherein the sample provides emission light that includes different first, second, third, and fourth spectral patterns in response to the first and second wavelengths;   a first detection camera configured to detect the first spectral pattern during a first measurement phase of the illumination/detection cycle, the first detection camera configured to detect the third spectral pattern during a second measurement phase of the illumination/detection cycle; and   a second detection camera configured to detect the second spectral pattern during the first measurement phase, the second detection camera configured to detect the fourth spectral pattern during the second measurement phase.   
     
     
         3 . The system of  claim 2 , wherein the first detection camera and the second detection camera concurrently detect the first and second spectral patterns during the first measurement phase. 
     
     
         4 . The system of  claim 2 , wherein the first detection camera successively detects the first and third spectral patterns. 
     
     
         5 . The system of  claim 2 , further comprising a detection assembly that includes the first and second detection cameras and a dichroic mirror, the dichroic mirror reflecting the first spectral pattern and the third spectral pattern during the first and second measurement phases, respectively, the dichroic mirror permitting the second spectral pattern and the fourth spectral pattern to pass therethrough during the first and second measurement phases, respectively. 
     
     
         6 . The system of  claim 5 , wherein the detection assembly includes a band pass filter, the first and third spectral patterns being directed along a reflection detection path toward the band pass filter prior to the first detection camera, the band pass filter configured to block high and low spectral content of at least one of the first or third spectral patterns. 
     
     
         7 . The system of  claim 6 , wherein the band pass filter has two pass bands that are separated from each other along a wavelength spectrum. 
     
     
         8 . The system of  claim 6 , wherein the detection assembly includes another band pass filter, the second and fourth spectral patterns being directed along a transmission detection path toward the other band pass filter prior to the second detection camera, the other band pass filter configured to block high and low spectral content of at least one of the second or fourth spectral patterns. 
     
     
         9 . The system of  claim 2 , wherein the first and second detection cameras output first, second, third and fourth data signals representative of detected portions of the first, second, third and fourth spectral patterns. 
     
     
         10 . The system of  claim 9 , wherein the illumination/detection cycle is repeated numerous times. 
     
     
         11 . The system of  claim 10 , further comprising a readout module that receives the first, second, third, and fourth data signals, the readout module configured to provide a plurality of images that are representative of the emission light detected by the first and second detection cameras. 
     
     
         12 . A method comprising:
 (a) exciting a sample with a first wavelength during a first excitation event, the sample providing first and second spectral patterns in response to the first wavelength;   (b) detecting the first spectral pattern with a first detection camera;   (c) detecting the second spectral pattern with a second detection camera;   (d) exciting the sample with a second wavelength during a second excitation event, the sample providing third and fourth spectral patterns in response to the second wavelength;   (e) detecting the third spectral pattern with the first detection camera; and   (f) detecting the fourth spectral pattern with the second detection camera.   
     
     
         13 . The method of  claim 12 , wherein detecting the first spectral pattern and detecting the second spectral pattern occurs concurrently. 
     
     
         14 . The method of  claim 12 , wherein detecting the first spectral pattern and detecting the third spectral pattern occurs separately. 
     
     
         15 . The method of  claim 12 , wherein the first spectral pattern is reflected by a dichroic member and the second spectral pattern is permitted to pass through the dichroic member and wherein the third spectral pattern is reflected by the dichroic member and the fourth spectral pattern is permitted to pass through the dichroic member. 
     
     
         16 . The method of  claim 15 , further comprising blocking high and low spectral content of at least one of the first or third spectral patterns. 
     
     
         17 . The method of  claim 16 , wherein a band pass filter blocks the high and the low spectral content, the band pass filter having two pass bands that are separated from each other along a wavelength spectrum. 
     
     
         18 . The method of  claim 12 , further comprising outputting first, second, third, and fourth data signals from the first and second detection cameras that are representative of detected portions of the first, second, third and fourth spectral patterns. 
     
     
         19 . The method of  claim 19 , wherein steps (a)-(f) constitute an illumination/detection cycle, the method further comprising repeating the illumination/detection cycle numerous times. 
     
     
         20 . The method of  claim 20 , further comprising providing a plurality of images that are representative of the emission light detected by the first and second detection cameras.