Automated system for two-dimensional electrophoresis

Abstract

The present invention provides an integrated, fully automated, high-throughput system for two-dimensional electrophoresis comprised of gel-making machines, gel processing machines, gel compositions and geometries, gel handling systems, sample preparation systems, software and methods. The system is capable of continuous operation at high-throughput to allow construction of large quantitative data sets.

What is claimed is:  
     
         1 . A computer-assisted method for selecting and directing the isolation of one or more biomolecules present in a two-dimensional array, comprising: 
 a purification step, wherein a plurality of biomolecules of interest are substantially isolated from a first biological sample;    a first separation step, wherein said biomolecules are separated according to a first physical or chemical property to form a one-dimensional array of biomolecules;    a second separation step, wherein said one-dimensional array of biomolecules is separated according to a second physical or chemical property to form said two-dimensional array;    imaging said two-dimensional array or a replica thereof to generate a computer-readable output comprising, for each of a plurality of biomolecules detected in said two-dimensional array, a pair of x,y coordinates and a signal value;    processing said output in at least one computer to select one or more of said detected biomolecules in accordance with previously ordained or operator-specified criteria; and optionally    generating machine-readable instructions that direct a robotic device to isolate at least one of said selected biomolecules from said two-dimensional array.    
     
     
         2 . The method according to    claim 1   , further comprising: 
 isolating at least one of said selected biomolecules from said two-dimensional array by means of said robotic device in accordance with said machine-readable instructions.    
     
     
         3 . The method according to    claim 1   , in which said biomolecules are proteins.  
     
     
         4 . The method according to    claim 1   , in which said two-dimensional array is contained in a polyacrylamide gel.  
     
     
         5 . The method according to    claim 4   , in which said biomolecules have been separated by isoelectric focusing, followed by electrophoresis in the presence of sodium dodecyl sulfate.  
     
     
         6 . The method according to    claim 4   , in which said polyacrylamide gel is bonded to a generally planar solid support such that the gel has two-dimensional spatial stability, and the support is substantially non-interfering with respect to detection of a detectable label carried by the proteins.  
     
     
         7 . The method according to    claim 6   , in which said polyacrylamide gel is covalently bonded to said solid support.  
     
     
         8 . The method according to    claim 6   , in which said detectable label is a fluorescent label.  
     
     
         9 . The method according to    claim 6   , in which said solid support is glass.  
     
     
         10 . A computer assisted method for excising a region of a gel containing desired biomolecules from a gel, comprising: 
 a) separating said desired biomolecules from undesired biomolecules in a 2-D electrophoresis method to yield a separation of biomolecules on said gel;    b) scanning said gel to yield a digitized image of biomolecules in said gel;    c) using data from said digitized image to position a cutter over said region of said gel containing said desired biomolecules wherein positioning of said cutter is computer controlled;    d) moving said cutter into said gel to cut said region of said gel containing said desired biomolecules; and    e) lifting said cutter to lift said region of said gel containing said desired biomolecules away from said gel.    
     
     
         11 . The method of    claim 10    wherein said biomolecules are selected from the group consisting of proteins, DNA and RNA.  
     
     
         12 . The method of    claim 10    wherein said biomolecules are radioactively labeled or fluorescently labeled.  
     
     
         13 . The method of    claim 10    wherein said biomolecules are stained prior to scanning.  
     
     
         14 . The method of    claim 13    wherein said stain is selected from the group consisting of Coomassie Brilliant Blue, a silver stain, a fluorescent stain and a negative stain.  
     
     
         15 . The method of    claim 10    wherein said 2-D electrophoresis method comprises a step of isoelectric focusing.  
     
     
         16 . The method of    claim 10    wherein said 2-D electrophoresis method comprises a step of SDS polyacrylamide gel electrophoresis.  
     
     
         17 . The method of    claim 10    wherein said 2-D electrophoresis method comprises a step of immobilized pH gradient (IPG) isoelectric focusing.  
     
     
         18 . The method of    claim 15    wherein said isoelectric focusing is performed on a gel covalently bonded to a solid support.  
     
     
         19 . The method of    claim 18    wherein said solid support is plastic or glass.  
     
     
         20 . The method of    claim 17    wherein said IPG isoelectric focusing is performed on a gel covalently bonded to a solid support.  
     
     
         21 . The method of    claim 20    wherein said solid support is plastic or glass.  
     
     
         22 . The method of    claim 10    wherein said gel is placed onto a supporting plate prior to moving said cutter into said gel.  
     
     
         23 . The method of    claim 10    wherein said scanning is performed by a CCD digitizer.  
     
     
         24 . The method of    claim 10    wherein said cutter is mounted on a movable, computer controlled X-Y frame.  
     
     
         25 . The method of    claim 24    wherein said cutter is suspended above and co-planar with said gel.  
     
     
         26 . The method of    claim 10    wherein more than one region of said gel is excised with each region being excised separately.  
     
     
         27 . A computer assisted method for excising a region of a gel containing desired biomolecules from a gel, comprising: 
 a) separating said desired biomolecules from undesired biomolecules in a 2-D electrophoresis method to yield a separation of biomolecules on said gel wherein said biomolecules are labeled to release radiation;    b) placing a film sensitive to said radiation in contact with or near said gel to expose said film and produce an exposed film;    c) developing said exposed film to produce a developed film;    d) scanning said developed film to yield a digitized image of biomolecules in said gel;    e) using data from said digitized image to position a cutter over said region of said gel containing said desired biomolecules wherein positioning of said cutter is computer controlled;    f) moving said cutter into said gel to cut said region of said gel containing said desired biomolecules; and    g) lifting said cutter to lift said region of said gel containing said desired biomolecules away from said gel.