System for cell-based screening

• US 20040009539A1
• Filed: 04/11/2003
• Published: 01/15/2004
• Est. Priority Date: 02/27/1997
• Status: Active Grant

• Current Assignee: CARL Zeiss JENA GmbH (Carl-Zeiss-Stiftung)
• Original Assignee: CARL Zeiss JENA GmbH (Carl-Zeiss-Stiftung)
• Inventors: Taylor, D., Dunlay, R.
• Granted Patent: US 7,235,373 B2
• Time in Patent Office: Days
• US Class Current: 435/7.2
• CPC Class Codes: B01J 2219/00315 Microtiter plates
• CPC Class Codes: B01J 2219/00317 Microwell devices, i.e. having large numbers of wells
• CPC Class Codes: B01J 2219/00707 separated from the reactor apparatus
• CPC Class Codes: B01J 2219/00743 Cells
• CPC Class Codes: B01L 3/5085 for multiple samples, e.g. microtitration plates
• CPC Class Codes: B82Y 30/00 Nanotechnology for materials or surface science, e.g. nanocomposites
• CPC Class Codes: C12Q 1/68 involving nucleic acids
• CPC Class Codes: C12Q 1/6813 Hybridisation assays
• CPC Class Codes: C12Q 2525/179 incorporating arbitrary or random nucleotide sequences
• CPC Class Codes: C40B 60/14 for creating libraries
• CPC Class Codes: G01N 15/1433 using image recognition
• CPC Class Codes: G01N 2015/1486 Counting the particles
• CPC Class Codes: G01N 2015/1497 Particle shape
• CPC Class Codes: G01N 2021/6419 Excitation at two or more wavelengths
• CPC Class Codes: G01N 2021/6482 Sample cells, cuvettes
• CPC Class Codes: G01N 21/6428 Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" in vivo A61B5/00; immunoassay G01N33/53
• CPC Class Codes: G01N 21/6452 Individual samples arranged in a regular 2D-array, e.g. multiwell plates
• CPC Class Codes: G01N 21/6458 Fluorescence microscopy fluorescence microscopes per se G02B21/0076 and G02B21/16
• CPC Class Codes: G01N 33/5005 involving human or animal cells immunoassay G01N33/56966; immunoassays of protozoa G01N33/56905; protozoa in screening assays C12Q1/025
• CPC Class Codes: G01N 33/5035 on sub-cellular localization
• CPC Class Codes: G01N 33/582 : with fluorescent label
• CPC Class Codes: G02B 21/16 : adapted for ultraviolet illumination ; Fluorescence microscopes G02B21/0076 takes precedence
• CPC Class Codes: G02B 21/365 : Control or image processing arrangements for digital or video microscopes G02B21/361, G02B21/362 take precedence
• CPC Class Codes: Y10S 436/80 : Fluorescent dyes, e.g. rhodamine
• CPC Class Codes: Y10S 436/807 : Apparatus included in process claim, e.g. physical support structures
• CPC Class Codes: Y10S 436/809 : Multifield plates or multicontainer arrays
• CPC Class Codes: Y10S 977/795 : Composed of biological material
• CPC Class Codes: Y10S 977/881 : Microscopy or spectroscopy, e.g. sem, tem
• CPC Class Codes: Y10S 977/92 : Detection of biochemical
• CPC Class Codes: Y10S 977/941 : including dna logic element
• CPC Class Codes: Y10S 977/942 : including protein logic element

Abstract

The invention relates to an optical system for determining the distribution, environment, or activity of fluorescently labeled reporter molecules in cells for the purpose of screening large numbers of compounds for specific biological activity. The invention involves providing cells containing fluorescent reporter molecules in an array of locations and scanning numerous cells in each location with a fluorescent microscope, 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.

Claims

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 2 or 3 different fluorescent reporter molecules are in the cell.

10. A cell screening system comprising:

(a) a fluorescent microscope having a microscope objective, 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 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.