Visual-servoing optical microscopy

• US 20040029213A1
• Filed: 06/26/2003
• Published: 02/12/2004
• Est. Priority Date: 06/08/2000
• Status: Active Grant

• Current Assignee: Regents of the University of California (University of California)
• Original Assignee: Regents of the University of California (University of California)
• Inventors: Parvin, Bahram, Callahan, Daniel E.
• Granted Patent: US 7,546,210 B2
• Time in Patent Office: Days
• US Class Current: 435/40.5
• CPC Class Codes: C12M 35/08 Chemical, biochemical or biological means, e.g. plasma jet, co-culture
• CPC Class Codes: C12M 41/46 of cellular or enzymatic activity or functionality, e.g. cell viability
• CPC Class Codes: G01N 15/1433 using image recognition
• CPC Class Codes: G06T 2207/10056 Microscopic image
• CPC Class Codes: G06T 2207/20081 Training; Learning
• CPC Class Codes: G06T 2207/30024 Cell structures in vitro; Tissue sections in vitro
• CPC Class Codes: G06T 7/0012 Biomedical image inspection
• CPC Class Codes: G06T 7/44 using image operators, e.g. filters, edge density metrics or local histograms
• CPC Class Codes: G06V 10/443 by matching or filtering
• CPC Class Codes: G06V 20/695 Preprocessing, e.g. image segmentation

Abstract

The present invention provides methods and devices for the knowledge-based discovery and optimization of differences between cell types. In particular, the present invention provides visual servoing optical microscopy, as well as analysis methods. The present invention provides means for the close monitoring of hundreds of individual, living cells over time: quantification of dynamic physiological responses in multiple channels; real-time digital image segmentation and analysis; intelligent, repetitive computer-applied cell stress and cell stimulation; and the ability to return to the same field of cells for long-term studies and observation. The present invention further provides means to optimize culture conditions for specific subpopulations of cells.

Claims

1. A method comprising:

(a) receiving cellular image data from a detection device that monitors cells or subcellular components of the cells;

(b) analyzing the cellular image data; and

(c) automatically actuating a plurality of stimulating devices adapted to stimulate the cells or subcellular components in response to the analyzed cellular image data.

2. The method of claim 1 wherein (c) comprises automatically and independently actuating the plurality of stimulating devices to stimulate the cells in response to the analyzed cellular image data.

3. The method of claim 1 wherein the analyzing the cellular image data comprises at least one analysis, selected from the group consisting of analyzing the morphometry of the cells, analyzing the color of the cells, and analyzing the motion of the cells.

4. The method of claim 1, wherein the cellular image data are compared with a reference database of cellular information.

5. The method of claim 1, wherein said cellular image data are compared with a remote reference database of cellular information.

6. The method of claim 1 wherein the detection device comprises a microscope and the method further comprises monitoring the cells using the VSOM.

7. The method of claim 1 wherein the stimulating devices are syringes.

8. The method of claim 1 further comprising:

(d) storing the cellular image data as a function of time.

9. The method of claim 1 wherein the cells are living and the subcellular components are in living cells.

10. The method of claim 1 further comprising:

(d) repeating steps (a) through (c).

11. A method of analyzing a cell array comprising the steps of:

a) providing;

i) a cell array, ii) an optical system, iii) a computing means, iv) at least on test stimulus, and iv) a robotic system;

b) detecting light from said cell said array by means of said optical system to provide a first signal;

c) providing said first signal corresponding to said light from said cell array to said computing means;

d) using said first signal to generate a second signal wherein said second signal controls the delivery of said at least one test stimulus to said cell array to produce a stimulated cell array;

e) and delivering said test stimulus, wherein said optical system further detects light emitted from said stimulated cell array.

12. The method of claim 11, wherein said test stimulus is automatically applied to said cell array according to said first signal.

13. The method of claim 11, wherein said robotic system delivers said test stimulus to said cell array through at least two microfluidic channels.

14. The method of claim 11, wherein said optical system comprises at least one microscope.

15. The method of claim 14, wherein said microscope is selected from the group consisting of confocal microscopes and fluorescence microscopes.

16. The method of claim 11, wherein said cell array comprises at least two cell types.

17. The method of claim 16, wherein said optical system scans said cell array to detect subcellular components within said cell types contained in said cell array.

18. The method of claim 17, wherein said subcellular components are selected from the group consisting of nuclei, nucleoli, mitochondria, lysosomes, phagolysosomes, and storage vesicles.

19. The method of claim 17, wherein at least one subcellular component is labelled.

20. The method of claim 18, wherein said subcellular component is labelled by said test stimulus.

21. The method of claim 17, wherein said subcellular component is labelled with at least one compound selected from the group consisting of fluorescent dyes, radioactive compounds, enzymes, and vital dyes.

22. The method of claim 11, wherein said optical system converts said label into digital data.

23. The method of claim 11, wherein said robotic system utilizes said digital data to automatically adjust the delivery of said biologically active ingredient to said cell array.

24. The method of claim 11, further comprising analyzing said light emitted from said stimulated cell array to produce a cellular information profile.

25. The method of claim 24, further comprising the step of comparing said cellular information profile with a reference database of cellular information.

26. A system for analyzing a cell array comprising at least one cell array, an optical system having light detection means for detecting light emitted from said cell array, a robotic system, and a computer system comprising a first program environment having means to accept input from said optical system and a second program environment having means to provide output to said robotic system.

27. The system of claim 26, wherein said robotic system further comprises at least two test stimulus sources.

28. The system of claim 26, wherein said at least two test stimulus sources comprise biologically active compositions.

29. The system of claim 26, wherein said at least two test stimulus sources are in fluidic communication with said at least one cell array.

30. The system of claim 26, wherein said cell array comprises at least two cell types.

31. The system of claim 26, further comprising a means for remote operation of said computer system.

32. The system of claim 26, wherein said optical system comprises a microscope.

33. A computer readable medium comprising:

(a) code for receiving cellular image data from a detection device that monitors cells or subcellular components;

(b) code for analyzing the cellular image data; and

(c) code for automatically adjusting a plurality of stimulating devices adapted to stimulate the cells or subcellular components in response to the analyzed cellular image data.

34. The computer readable medium of claim 33, wherein the computer readable medium further comprises:

(d) code for controlling a visual servoing optical microscope (VSOM) monitoring the cells.

35. The computer readable medium of claim 33, wherein the computer readable medium further comprises at least one of

(d) code for analyzing the size of the cells, code for analyzing the color of the cells, and code for analyzing the motion of the cells.

36. The computer readable medium of claim 33, further comprising:

(d) code for automatically and independently actuating the plurality of stimulating devices to stimulate the cells in response to the analyzed cellular image data.

37. The computer readable medium of claim 33, wherein the stimulating devices are syringes.

38. The computer readable medium of claim 33, further comprising code for storing the cellular image data as a function of time.

39. The computer readable medium of claim 33, wherein the cells are living and the subcellular components are in living cells.