Methods and apparatus for simultaneously detecting a large range of protein concentrations

Abstract

Some embodiments described herein relate to a method that includes separating an analyte-containing sample via electrophoresis in a capillary. The capillary is loaded with a chemiluminescence agent, such as luminol, that is configured to react with the analyte (e.g., HRP-conjugated proteins) to produce a signal indicative of a concentration and/or quantity of analyte at each location along the length of the capillary. A first image of the capillary containing the analytes and the chemiluminescence agent is captured over a first period of time. A second image of the capillary containing the analytes and the chemiluminescence agent is captured over a second, longer, period of time. A concentration and/or quantity of a first population of analytes at a first location is determined using the first image, and a concentration and/or quantity of a second population of analytes at a second location is determined using the second image.

What is claimed is: 
     
         1 . A method, comprising:
 separating a sample containing a first population of proteins and a second population of proteins via capillary electrophoresis such that the first population of proteins is located at a first position along a length of a capillary and the second population of proteins is located at a second position along the length of the capillary;   capturing a plurality of images of the capillary, each image from the plurality of images having a different exposure period;   selecting a first image from the plurality of images in which a strength of a first optical signal indicative of a quantity of the first population of proteins exceeds a predetermined threshold, the first optical signal changing with time;   determining an initial intensity of the first optical signal by dividing the strength of the first optical signal by an exposure period for the first image;   calculating a quantity of the first population of proteins based on the initial intensity of the first optical signal;   selecting a second image from the plurality of images in which a strength of a second optical signal indicative of a quantity of the second population of proteins exceeds the predetermined threshold, the second optical signal changing with time; and   calculating a quantity of the second population of proteins based on the second optical signal and an exposure period for the second image.   
     
     
         2 . The method of  claim 1 , wherein a quantity of the first population of proteins is at least three times greater than a quantity of the second population of proteins. 
     
     
         3 . The method of  claim 1 , wherein a quantity of the first population of proteins is at least ten times greater than a quantity of the second population of proteins. 
     
     
         4 . The method of  claim 1 , further comprising:
 determining an initial intensity of the second optical signal by dividing the strength of the second optical signal by the exposure period for the second image, the quantity of the second population calculated based on the initial intensity of the second optical signal.   
     
     
         5 . The method of  claim 1 , wherein the plurality of images is divided into subsets each having a nominal exposure time, an exposure time of each image from each subset varying by 50% or less from the nominal exposure time for that subset. 
     
     
         6 . The method of  claim 1 , wherein each subset of images from the plurality of images has a nominal exposure time of twice a nominal exposure time of an immediately prior subset of images from the plurality of images. 
     
     
         7 . The method of  claim 1 , wherein each image from the plurality of images has an exposure time of twice an exposure time of an immediately prior image from the plurality of images. 
     
     
         8 . The method of  claim 1 , further comprising:
 adding one of a chemiluminescence agent or a detection agent to the capillary prior to capturing an image from the plurality of images.   
     
     
         9 . The method of  claim 1 , further comprising:
 adding one of a chemiluminescence agent or a chromogenic agent to the capillary prior to capturing each image from the plurality of images.   
     
     
         10 . The method of  claim 1 , wherein the first optical signal and the second optical signal are each associated with a detection agent, the detection agent at the second position depleted by during the exposure period for the first image. 
     
     
         11 . The method of  claim 1 , wherein the second optical signal is below a detection threshold in the first image. 
     
     
         12 . The method of  claim 1 , wherein the first position of the capillary and the second position of the capillary both appear in each of the first image and the second image. 
     
     
         13 . The method of  claim 1 , wherein:
 the second image is captured after the first image; and   the exposure period for the second image is longer than the exposure period for the first image.   
     
     
         14 . The method of  claim 1 , wherein the plurality of images is divided into subsets of images, each subset of images including a plurality of images sharing an exposure time that is different from each other subset of images. 
     
     
         15 . The method of  claim 1 , wherein:
 the plurality of images includes a first subset of images and a second subset of images, each image from the first subset of images having a first exposure time and each image from the second subset of images having a second exposure time, the first subset of images captured before the second subset of images.   
     
     
         16 . A non-transitory processor-readable medium storing code, the code comprising instructions to cause a processor to:
 send a signal to a voltage source to cause the voltage source to apply an electric potential across a capillary to cause a sample containing a first population of proteins and a second population of proteins to be separated via capillary electrophoresis such that the first population of proteins is located at a first position along a length of a capillary and the second population of proteins is located at a second position along the length of the capillary;   receive, from a detector, a plurality of images of the capillary, each image from the plurality of images having a different exposure period;   select a first image from the plurality of images in which a strength of a first optical signal indicative of a quantity of the first population of proteins exceeds a predetermined threshold, the first optical signal changing with time;   determine an initial intensity of the first optical signal by dividing the strength of the first optical signal by an exposure period for the first image;   calculate a quantity of the first population of proteins based on the initial intensity of the first optical signal;   select a second image from the plurality of images in which a strength of a second optical signal indicative of a quantity of the second population of proteins exceeds the predetermined threshold, the second optical signal changing with time; and   calculate a quantity of the second population of proteins based on the second optical signal and an exposure period for the second image.   
     
     
         17 . The non-transitory processor-readable medium of  claim 16 , the code further comprising instructions to cause the processor to:
 determine an initial intensity of the second optical signal by dividing the strength of the second optical signal by the second duration, the quantity of the second population of proteins calculated based on the initial intensity of the second optical signal.   
     
     
         18 . The non-transitory processor-readable medium of  claim 16 , the code further comprising instructions to cause the processor to:
 send a signal to cause one of a chemiluminescence agent or a detection agent to be added to the capillary prior to capturing an image from the plurality of images.   
     
     
         19 . The non-transitory processor-readable medium of  claim 16 , the code further comprising instructions to cause the processor to:
 send a signal to cause one of a chemiluminescence agent or a detection agent to be added to the capillary prior to capturing each image from the plurality of images.   
     
     
         20 . The non-transitory processor-readable medium of  claim 16 , wherein:
 the second image is captured after the first image; and   the exposure period for the second image is longer than the exposure period for the first image.