System for cell-based screening

Abstract

The present invention provides systems, methods, and screens for an optical system analysis of cells to rapidly determine the distribution, environment, or activity of fluorescently labeled reporter molecules in cells for the purpose of screening large numbers of compounds for those that specifically affect particular biological functions. The invention involves providing cells containing fluorescent reporter molecules in an array of locations and scanning numerous cells in each location with a high magnification fluorescence optical system, converting the optical information into digital data, and utilizing the digital data to determine the distribution, environment or activity of the fluorescently labeled reporter molecules in the cells. The array of locations may be an industry standard 96 well or 384 well microtiter plate or a microplate which is a microplate having a cells in a micropaterned array of locations. The invention includes apparatus and computerized method for processing, displaying and storing the data.

What is claimed is:  
     
         1 . A method for analyzing cells comprising: 
 (a) providing an array of locations which contain multiple cells wherein the cells contain one or more fluorescent reporter molecules;    (b) scanning multiple cells in each of the locations containing cells to obtain fluorescent signals from the fluorescent reporter molecule in the cells;    (c) converting the fluorescent signals into digital data; and    (d) utilizing the digital data to determine the distribution, environment or activity of the fluorescent reporter molecules within the cells.    
     
     
         2 . The method of  claim 1  wherein the array of locations are wells in a microtiter plate.  
     
     
         3 . The method of  claim 1  wherein the array of locations are microwells on a microplate.  
     
     
         4 . The method of  claim 1  wherein the fluorescent reporter is added to the cell.  
     
     
         5 . The method of  claim 1  wherein the fluorescent reporter is produced by the cell.  
     
     
         6 . The method of  claim 1  wherein a computer means converts the digital data into the difference between the average cytoplasmic reporter molecule fluorescent intensity and the average nucleus fluorescent reporter molecule intensity.  
     
     
         7 . The method of  claim 1  wherein a computer means converts the digital data into the average cytoplasmic fluorescent reporter molecule intensity within the nucleus region.  
     
     
         8 . The method of  claim 1  wherein a computer means converts the digital data into the average fluorescent reporter molecule intensity within the cytoplasmic mask.  
     
     
         9 . The method of  claim 1  wherein multiple different fluorescent reporter molecules are in the cell.  
     
     
         10 . A cell screening system comprising: 
 (a) a high magnification fluorescence optical system having an objective lens, an XY stage adapted for holding a plate with an array of locations for holding cells and having a means for moving the plate to align the locations with the microscope objective and a means for moving the plate in the direction to effect focusing;    (b) a digital camera;    (c) a light source having optical means for directing excitation light to cells in the array locations and a means for directing fluorescent light emitted from the cells to the digital camera; and    (d) a computer means for receiving and processing digital data from the digital camera wherein the computer means includes: 
 i) a digital frame grabber for receiving the images from the camera,  
 ii) a display for user interaction and display of assay results,  
 iii) digital storage media for data storage and archiving, and  
 iv) means for control, acquisition, processing and display of results.  
   
     
     
         11 . The cell screening system of  claim 10  having a PC screen operatively associated with the computer means, for displaying graphs of data and images of cells having fluorescent reporter molecules.  
     
     
         12 . The cell screening system of  claim 10  wherein the computer means stores the data in a bioinformatics data base.  
     
     
         13 . The method of  claim 1  further comprising scanning multiple cells in each of the array of locations containing cells in high throughput mode, and selectively scanning only a subset of the locations containing cells in a high content mode.  
     
     
         14 . The cell screening system of  claim 10  further comprising an attached reader which measures a signal from the array of locations which contain multiple cells, and a method to transfer both the array of locations which contain multiple cells and the measurements to the cell screening system.  
     
     
         15 . The cell screening system of  claim 10  wherein the optical means comprise a mechanical-optical means for changing the magnification of the system.  
     
     
         16 . The cell screening system of  claim 10  further comprising a chamber and control system to maintain the temperature, CO 2  concentration and humidity surrounding the array of ocations which contain multiple cells.  
     
     
         17 . The cell screening system of  claim 10  wherein the optical means comprises a confocal scanning illumination and detection system.  
     
     
         18 . A machine readable storage medium comprising a program containing a set of instructions for causing a cell screening system to execute the procedures set forth in FIG. 9, wherein the cell screening system comprises a high magnification fluorescence optical system with a stage adapted for holding cells and a means for moving the stage, a digital camera, a light source for receiving and processing the digital data from the digital camera, and a computer means for receiving and processing the digital data from the digital camera.  
     
     
         19 . A machine readable storage medium comprising a program containing a set of instructions for causing a cell screening system to execute the procedures set forth in FIG. 11, wherein the cell screening system comprises a high magnification fluorescence optical system with a stage adapted for holding cells and a means for moving the stage, a digital camera, a light source for receiving and processing the digital data from the digital camera, and a computer means for receiving and processing the digital data from the digital camera.  
     
     
         20 . A machine readable storage medium comprising a program containing a set of instructions for causing a cell screening system to execute the procedures set forth in FIG. 12, wherein the cell screening system comprises a high magnification fluorescence optical system with a stage adapted for holding cells and a means for moving the stage, a digital camera, a light source for receiving and processing the digital data from the digital camera, and a computer means for receiving and processing the digital data from the digital camera.  
     
     
         21 . A machine readable storage medium comprising a program containing a set of instructions for causing a cell screening system to execute the procedures set forth in FIG. 13, wherein the cell screening system comprises a high magnification fluorescence optical system with a stage adapted for holding cells and a means for moving the stage, a digital camera, a light source for receiving and processing the digital data from the digital camera, and a computer means for receiving and processing the digital data from the digital camera.  
     
     
         22 . A machine readable storage medium comprising a program containing a set of instructions for causing a cell screening system to execute the procedures set forth in FIG. 14, wherein the cell screening system comprises a high magnification fluorescence optical system with a stage adapted for holding cells and a means for moving the stage, a digital camera, a light source for receiving and processing the digital data from the digital camera, and a computer means for receiving and processing the digital data from the digital camera.  
     
     
         23 . A machine readable storage medium comprising a program containing a set of instructions for causing a cell screening system to execute the procedures set forth in FIG. 15, wherein the cell screening system comprises a high magnification fluorescence optical system with a stage adapted for holding cells and a means for moving the stage, a digital camera, a light source for receiving and processing the digital data from the digital camera, and a computer means for receiving and processing the digital data from the digital camera.  
     
     
         24 . A machine readable storage medium comprising a program containing a set of instructions for causing a cell screening system to execute procedures for detecting the distribution and activity of specific cellular constituents and processes, wherein the cell screening system comprises a high magnification fluorescence optical system with a stage adapted for holding cells and a means for moving the stage, a digital camera, a light source for receiving and processing the digital data from the digital camera, and a computer means for receiving and processing the digital data from the digital camera.  
     
     
         25 . The machine readable storage medium of  claim 24 , wherein the specific cellular process comprises the nuclear translocation of a protein.  
     
     
         26 . The machine readable storage medium of  claim 25  wherein said protein comprises a membrane protein.  
     
     
         27 . The machine readable storage medium of  claim 24 , wherein the specific cellular process comprises cellular hypertrophy.  
     
     
         28 . The machine readable storage medium of  claim 24 , wherein the specific cellular process comprises apoptosis.  
     
     
         29 . The machine readable storage medium of  claim 24 , wherein the specific cellular process comprises protease-induced translocation of a protein.  
     
     
         30 . A machine readable storage medium comprising a program containing a set of instructions for causing a cell screening system to execute procedures for identifying novel receptor agonists and antagonists, wherein the cell screening system comprises a high magnification fluorescence optical system with a stage adapted for holding cells and a means for moving the stage, a digital camera, a light source for receiving and processing the digital data from the digital camera, and a computer means for receiving and processing the digital data from the digital camera.  
     
     
         31 . A machine readable storage medium comprising a program containing a set of instructions for causing the cell screening system of  claim 10  to execute the procedures set forth in FIG. 9.  
     
     
         32 . A machine readable storage medium comprising a program containing a set of instructions for causing the cell screening system of  claim 10  to execute the procedures set forth in FIG. 11.  
     
     
         33 . A machine readable storage medium comprising a program containing a set of instructions for causing the cell screening system of  claim 10  to execute the procedures set forth in FIG. 12.  
     
     
         34 . A machine readable storage medium comprising a program containing a set of instructions for causing the cell screening system of  claim 10  to execute the procedures set forth in FIG. 13.  
     
     
         35 . A machine readable storage medium comprising a program containing a set of instructions for causing the cell screening system of  claim 10  to execute the procedures set forth in FIG. 14.  
     
     
         36 . A machine readable storage medium comprising a program containing a set of instructions for causing the cell screening system of  claim 10  to execute the procedures set forth in FIG. 15.  
     
     
         37 . A machine readable storage medium comprising a program containing a set of instructions for causing the cell screening system of  claim 10  to execute procedures for detecting the distribution and activity of specific cellular constituents and processes.  
     
     
         38 . The machine readable storage medium of  claim 37 , wherein the specific cellular process comprises the nuclear translocation of a protein.  
     
     
         39 . The machine readable storage medium of  claim 37  wherein said protein comprises a membrane protein.  
     
     
         40 . The machine readable storage medium of  claim 37 , wherein the specific cellular process comprises cellular hypertrophy.  
     
     
         41 . The machine readable storage medium of  claim 37 , wherein the specific cellular process comprises apoptosis.  
     
     
         42 . The machine readable storage medium of  claim 37 , wherein the specific cellular process comprises protease-induced translocation of a protein.  
     
     
         43 . A machine readable storage medium comprising a program containing a set of instructions for causing the cell screening system of  claim 10  to execute procedures for identifying novel receptor agonists and antagonists, wherein the cell screening system comprises a high magnification fluorescence optical system with a stage adapted for holding cells and a means for moving the stage, a digital camera, a light source for receiving and processing the digital data from the digital camera, and a computer means for receiving and processing the digital data from the digital camera.