OPTICAL CYTOMETRY

• US 20100284016A1
• Filed: 05/06/2009
• Published: 11/11/2010
• Est. Priority Date: 05/06/2009
• 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: Reed, Jason C., Gimzewski, James K., Teitell, Michael A.
• Granted Patent: US 8,599,383 B2
• Time in Patent Office: Days
• US Class Current: 356/451
• CPC Class Codes: G01J 3/453 by correlation of the amplitudes
• CPC Class Codes: G01N 33/5008 for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics
• CPC Class Codes: G02B 21/0056 based on optical coherence, e.g. phase-contrast arrangements, interference arrangements
• CPC Class Codes: G02B 21/32 Micromanipulators structurally combined with microscopes
• CPC Class Codes: G02B 5/126 including curved refracting surface

Abstract

The present invention provides optical systems and methods for determining a characteristic of a cell, such as cell type, cellular response to a biochemical event, biological state and the like. The methods typically involve using interferometry to observe membrane properties in a cell and then use this information to determine one or more characteristics of a cell. The methods of the invention are useful for applications such as drug screening as well as diagnostic techniques.

Claims

1. A system for obtaining an image of a cell comprising:

(a) a microscope capable of measuring a feature of interest in a sample.

(b) a detector operatively coupled to the microscope;

(c) a sample assembly comprising an observation chamber adapted to contain the cell; and

(d) a plurality of reflective microparticles capable of adhering to the cell, wherein the average diameter of the reflective microparticles is between 0.5 μ

m and 30 μ

m.

2. The system of claim 1, wherein the microscope is a confocal microscope

3. The system of claim 1, wherein the microscope is an interference microscope capable of observing interference fringes and the system further comprises a reference assembly adapted to substantially match an optical path length of the sample assembly.

4. The system of claim 3, wherein the reference assembly further comprises:

a first optical window;

a first housing element adapted to hold the first optical window;

a second optical window;

a second housing element adapted to hold the second optical window; and

a plurality of spherical spacer elements disposable between the first optical window and the second optical window and adapted to separate the first and second optical windows to a defined distance.

5. The system of claim 4, wherein the sample assembly further comprises:

a viewing window;

a first housing element adapted to hold the viewing window;

and wherein the thickness of the viewing window is equivalent to the combined thickness of the first and second optical windows in the reference assembly.

6. The system of claim 1, wherein the observation chamber comprises at least one perfusion conduit adapted to circulate a cell media within the chamber.

7. The system of claim 1, wherein the reflective microparticles are spherical magnetic microparticles having an average diameter of between 1 μ

m and 15 μ

m.

8. The system of claim 1, wherein the reflective microparticles comprise a gradient index (GRIN) spherical lens.

9. The system of claim 1, further comprising a magnet disposed below the observation chamber and oriented coaxially with an optical axis.

10. The system of claim 9, wherein the magnet is operatively coupled to a motorized micrometer and adapted to exert a magnetic force of between 0 Newtons and 5 nanoNewtons on a magnetic reflective microparticle adhered to the surface of the cell.

11. The system of claim 9, wherein the magnet is adapted to generate a magnetic field of between 200 Gauss and 3 kiloGauss and/or a magnetic field gradient range of between 300,000 to 800,000 Gauss/meter.

12. The system of claim 9, wherein magnet is operatively coupled to the system so as to allow the measurement of an axial position of a magnetic reflective microparticle in a sample.

13. The system of claim 9, wherein magnet is operatively coupled to the motorized micrometer configured to allow the measurement of a z motion of a magnetic reflective microparticle in a sample.

14. The system of claim 1, further comprising a processor element and a memory storage element adapted to process and store one or more images of the cell.

15. A method for observing a property of a cell, the method comprising:

(a) adhering a reflective magnetic microparticle to the cell;

(b) placing the cell in a cell observation chamber of a microscope;

(c) exposing the cell coated with the microparticle to a magnetic field; and

(d) using the microscope to observe the movement of the microparticle adhered to the cell in response to the applied magnetic field, wherein the movement of the reflective microparticle adhered to the cell correlates to a property of the cell, so that a property of the cell is observed.

16. The method of claim 15, wherein the method comprises measuring:

an axial position of a magnetic reflective microparticle in a sample comprising the cell;

or

a z motion of a magnetic reflective microparticle in a sample comprising the cell.

17. The method of claim 15, wherein the microscope is confocal microscope.

18. The method of claim 15, wherein the microscope is an interference microscope capable of observing interference fringes.

19. The method of claim 18, wherein the method is performed using a system comprising:

(a) a detector operatively coupled to the microscope;

(b) a sample assembly comprising an observation chamber adapted to contain the cell;

(c) a reference assembly comprising a reference chamber adapted to contain a fluid;

(d) a memory storage element adapted to store one or more images of the cell; and

(e) a processor element adapted to process one or more images of the cell.

20. The method of claim 18, wherein the method is used to observe an optical thickness or a cell mass property of a live cell in an aqueous medium.

21. The method of claim 18, wherein the method is used to observe a viscoelastic property of a live cell in an aqueous medium.

22. The method of claim 18, wherein the method is used to observe a population of live cells.

23. The method of claim 22, wherein the method is used to observe resting and dynamic responses to stimuli in a population of live cells.

24. The method of claims 18, wherein the property is observed in response to the cell'"'"'s exposure to:

the magnetic field;

or

a composition introduced into the cell'"'"'s media.

25. The method of claim 15, further comprising removing the cell from the observation chamber and manipulating the cell for a further analysis.

26. The method of claim 19, wherein the method is used to obtain information comprising a cell specific profile of a live cell in an aqueous medium and to store this information in the memory storage element.

27. A system for observing a property of a deformable material comprising:

(a) a microscope capable of measuring a feature of interest in a sample;

(b) a detector operatively coupled to the microscope;

(c) a sample assembly comprising an observation chamber adapted to contain the deformable material; and

(d) a plurality of reflective microparticles capable of adhering to the deformable material, wherein the average diameter of the reflective microparticles is between 0.5 μ

m and 30 μ

m.

28. A method for observing a property of a deformable material comprising using the system of claim 27 in one or more optical profiling techniques comprising:

confocal holography;

digital holography;

spectrally resolved interferometry;

or

wavelength scanning interferometry.

29. A reflective microparticle comprising a gradient index (GRIN) spherical lens.

30. The reflective microparticle of claim 29, wherein the microparticle is coupled to a flexible tether.

31. The reflective microparticle of claim 29, wherein the microparticle is coupled to an optical fiber.

32. The reflective microparticle of claim 29, wherein the microparticle is operatively coupled to an endoscope.

33. The reflective microparticle of claim 29, wherein the microparticle comprises a plurality of material layers, wherein refractive indices of the material layers decrease from the center of the microparticle.

34. A system for obtaining an image of a cell comprising:

(a) a interference microscope capable of extracting information from interferometric fringes;

(b) a detector operatively coupled to the interference microscope;

(c) a sample assembly comprising an observation chamber adapted to contain the cell,

(d) a reference assembly adapted to substantially match an optical path length of the sample assembly, and

(e) a plurality of reflective microparticles capable of adhering to the cell, wherein the average diameter of the reflective microparticles is between 0.5 mm and 30 mm.

35. A method for observing a property of a deformable material, the method comprising:

(a) adhering a reflective magnetic microparticle to the deformable material;

(b) placing the deformable material in a observation chamber of a interference microscope capable of extracting information from interferometric fringes;

(c) exposing the deformable material coated with the microparticle to a magnetic field; and

(d) using the microscope to observe the movement of the microparticle adhered to the deformable material in response to the applied magnetic field, wherein the movement of the reflective microparticle adhered to the deformable material correlates to a property of the deformable material, so that a property of the deformable material is observed.

36. A method for observing a property of a cell, the method comprising:

(a) adhering a reflective magnetic microparticle to the cell;

(b) placing the cell in a observation chamber of a interference microscope capable of extracting information from interferometric fringes;

(c) exposing the cell coated with the microparticle to a magnetic field; and

(d) using the microscope to observe the movement of the microparticle adhered to the cell in response to the applied magnetic field, wherein the movement of the reflective microparticle adhered to the cell correlates to a property of the cell, so that a property of the cell is observed.

37. A system for observing a property of a deformable material comprising:

(a) a microscope capable of measuring a feature of interest in a sample;

(b) a detector operatively coupled to the microscope;

(c) a sample assembly adapted to contain the deformable material;

(d) a magnetic microparticle adhered to the deformable material; and

(e) a magnetic field exerting a magnetic force on the magnetic microparticle.

38. A system for observing a property of a deformable material comprising:

(a) an interference microscope capable of extracting information from interferometric fringes;

(b) a detector operatively coupled to the interference microscope;

(c) a sample assembly adapted to contain the deformable material;

(d) a reference assembly in the interference microscope adapted to substantially match an optical path length of the sample assembly;

(e) a magnetic microparticle adhered to the deformable material; and

(f) a magnetic field exerting a magnetic force on the magnetic microparticle.