Biosensor array comprising cell populations confined to microcavities

• US 6,377,721 B1
• Filed: 03/02/1999
• Issued: 04/23/2002
• Est. Priority Date: 03/02/1998
• Status: Expired due to Term

• Current Assignee: Trustees of Tufts College
• Original Assignee: Trustees of Tufts College
• Inventors: Taylor, Laura C., Walt, David R.
• Primary Examiner(s): Healy, Brian
• Application Number:
  US09/260,963
• Time in Patent Office: 1,148 Days
• Field of Search: 385/12, 385/13, 385/31, 385/33, 385/38, 385/147, 385/901, 385/14, 385/115, 385/117, 385/116, 385/120, 435/808, 359/900, 250/556, 436/164, 436/172, 436/800, 436/805
• US Class Current: 385/12
• CPC Class Codes: B01J 2219/00317 Microwell devices, i.e. having large numbers of wells
• CPC Class Codes: B01J 2219/005 Beads
• CPC Class Codes: B01J 2219/00511 Walls of reactor vessels
• CPC Class Codes: B01J 2219/00524 in the shape of fiber bundles
• CPC Class Codes: B01J 2219/00585 Parallel processes
• CPC Class Codes: B01J 2219/00596 Solid-phase processes
• CPC Class Codes: B01J 2219/00605 the compounds being directly bound or immobilised to solid supports
• CPC Class Codes: B01J 2219/00612 the surface being inorganic
• CPC Class Codes: B01J 2219/00621 by physical means, e.g. trenches, raised areas
• CPC Class Codes: B01J 2219/00626 Covalent
• CPC Class Codes: B01J 2219/00628 Ionic
• CPC Class Codes: B01J 2219/0063 Other, e.g. van der Waals forces, hydrogen bonding
• CPC Class Codes: B01J 2219/00637 by coating it with another layer
• CPC Class Codes: B01J 2219/00648 by the use of solid beads
• CPC Class Codes: B01J 2219/00659 Two-dimensional arrays
• CPC Class Codes: B01J 2219/00704 integrated with the reactor apparatus
• CPC Class Codes: B01L 2200/0668 Trapping microscopic beads
• CPC Class Codes: B01L 2300/0654 Lenses; Optical fibres
• CPC Class Codes: B01L 2300/0819 Microarrays; Biochips
• CPC Class Codes: B01L 3/5085 for multiple samples, e.g. microtitration plates
• CPC Class Codes: G01N 21/253 : for batch operation, i.e. multisample apparatus analytical automats G01N35/00
• CPC Class Codes: G01N 33/54373 : involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings
• CPC Class Codes: Y10S 385/901 : Illuminating or display apparatus
• CPC Class Codes: Y10S 435/808 : Optical sensing apparatus
• CPC Class Codes: Y10S 436/80 : Fluorescent dyes, e.g. rhodamine
• CPC Class Codes: Y10S 436/805 : Optical property

Abstract

A biosensor, sensor array, sensing method and sensing apparatus are provided in which individual cells or randomly mixed populations of cells, having unique response characteristics to chemical and biological materials, are deployed in a plurality of discrete sites on a substrate. In a preferred embodiment, the discrete sites comprise microwells formed at the distal end of individual fibers within a fiber optic array. The biosensor array utilizes an optically interrogatable encoding scheme for determining the identity and location of each cell type in the array and provides for simultaneous measurements of large numbers of individual cell responses to target analytes. The sensing method utilizes the unique ability of cell populations to respond to biologically significant compounds in a characteristic and detectable manner. The biosensor array and measurement method may be employed in the study of biologically active materials, in situ environmental monitoring, monitoring of a variety of bioprocesses, and for high throughput screening of large combinatorial chemical libraries.

Claims

1. A biosensor for detecting the response of individual cells to at least one analyte of interest comprising:

a) a substrate comprising a plurality of wells; and

b) a population of cells comprising at least a first and second subpopulation, each cell dispersed into a different well, wherein each subpopulation is encoded with at least one optically interrogatable material.

3. The biosensor of claim 1 or claim 2 wherein said wells have an internal diameter ranging from about 1 um to about 200 um, a depth ranging from about 0.25 um to about 200 um, and a volume ranging from about 1 fL to about 5 nanoliters.

5. The biosensor of claim 1 or 2 wherein said wells are coated with a biologically compatible material.

6. The biosensor of claim 1 or 2 wherein an indicator compound is attached to or inserted into at least one of said cells.

21. The biosensor of claim 1 or claim 2 wherein said cells are encoded by attaching said optically interrogatable material to said cells.

22. The biosensor of claim 1 or claim 2 wherein said cells uptake said optically interrogatable material.

23. The biosensor of claim 1 or claim 2 wherein said cells are encoded by injecting said optically interrogatable material.

24. The biosensor of claim 1 or claim 2 wherein said cells are genetically engineered to exhibit chemiluminescence.

25. The biosensor of claim 1 or claim 2 wherein said cells are genetically engineered to exhibit bioluminescence.

26. The biosensor of claim 1 or claim 2 wherein said optically interrogatable material is a dye.

27. The biosensor of claim 1 or claim 2 wherein said optically interrogatable material is a chromophore.

28. The biosensor of claim 1 or claim 2 wherein said optically interrogatable material is a fluorophore.

29. The biosensor of claim 1 or claim 2 wherein said optically interrogatable material is a chemiluminescent compound.

30. The biosensor of claim 1 or claim 2 wherein said optically interrogatable material is a bioluminescent compound.

2. A biosensor for detecting the response of individual cells to at least one analyte of interest comprising:

a) a fiber optic bundle comprising a plurality of fibers, said fiber optic bundle comprising at least a first surface comprising wells; and

b) a plurality of cells, each of said cells distributed into a different well, wherein each cell is encoded with at least one optically interrogatable material.

4. The biosensor of claim 2 wherein said fibers further comprise a fiber core having a diameter ranging from about 1 um to about 200 um.

7. The biosensor of claim 2 wherein said plurality of cells comprises at least two cell populations.

8. An apparatus for detecting the response of individual cells to at least one analyte of interest comprising:

a) a biosensor array comprising;

i) a substrate comprising a plurality of wells; and

ii) a population of cells comprising at least a first and second subpopulation, each cell dispersed into a different well, wherein each subpopulation is encoded with at least one optically interrogatable material; and

b) a detector optically coupled to and in optical communication with said wells on said substrate, said detector capable of detecting an optical response of said cells dispersed in said wells to an analyte.

11. An apparatus according to claim 8 further comprising an image capturing device for capturing images of detected optical responses from a plurality of said cells.

12. The apparatus of claim 11 wherein said image capturing device comprises a charge coupled device or CCD camera.

9. An apparatus for detecting the response of individual cells to at least one analyte of interest comprising:

a) a biosensor comprising;

i) a fiber optic bundle comprising a plurality of fibers, each fiber comprising at least a first surface comprising wells; and

ii) a plurality of cells, each of said cells distributed into a different well, wherein each cell is encoded with at least one optically interrogatable material; and

b) a detector.

10. An apparatus according to claim 9 further comprising an excitation light energy device coupled to an end of said fibers.

13. A method for detecting the response of individual cells to at least one analyte of interest comprising:

a) providing a biosensor array comprising;

i) a substrate comprising a plurality of wells; and

ii) a population of cells comprising at least a first and second subpopulation, each cell dispersed into a different well, wherein each subpopulation is encoded with at least one different optically interrogatable material;

b) contacting said biosensor array with an analyte of interest; and

c) detected an optical response of said cells.

14. A method according to claim 13 wherein said analyte of interest comprises an optically detectable label.

15. A method according to claim 13 wherein said biosensor array is contacted with a plurality of analytes.

16. A method of making a biosensor array for detecting the response of individual cells to at least one analyte of interest comprising:

a) providing a substrate comprising a fiber optic array, said fiber optic array comprising;

i) a plurality of fibers, each fiber comprising at least a first surface; and

ii) a plurality of wells each formed on said first surface; and

b) contacting said first surface of said fiber optic array with a plurality of cells such that each cell is inserted into one of said wells.

17. A method according to claim 16 wherein said plurality of cells comprises at least two cell populations.

18. A method according to claim 17 wherein each cell population is encoded with at least one optically interrogatable material selected from the group consisting of a dye, a fluorophore, a chromophore, a chemiluminescent compound and a bioluminescent compound.

19. A method according to claim 18 further comprising indexing the location of individual cells in said array by emitted light energy produced by illuminating said encoded cells with excitation light energy.

20. A method for detecting the response of individual cells to at least one analyte of interest comprising:

a) providing a biosensor comprising;

i) a fiber optic bundle comprising a plurality of fibers, each fiber comprising at least a first surface comprising wells;

ii) a plurality of cells, each of said cells distributed into a different well, wherein each cell is encoded with at least one optically interrogatable material;

b) contacting said biosensor with an analyte of interest; and

c) detecting an optical response of said cells.