Binding method and apparatus for sorting objects

• US 8,067,170 B2
• Filed: 09/11/2008
• Issued: 11/29/2011
• Est. Priority Date: 09/11/2007
• Status: Expired due to Fees

• Current Assignee: Arryx, Inc. (Haemonetics Corporation)
• Original Assignee: Arryx, Inc. (Haemonetics Corporation)
• Inventors: Chakrabarty, Tania
• Primary Examiner(s): Babic; Christopher M.
• Application Number:
  US12/232,164
• Publication Number:
  US 20090068671A1
• Time in Patent Office: 1,174 Days
• Field of Search: None
• US Class Current: 435/6.1
• CPC Class Codes: G01N 2800/367 Infertility, e.g. sperm disorder, ovulatory dysfunction
• CPC Class Codes: G01N 33/5091 for testing the pathological state of an organism
• CPC Class Codes: G01N 33/56966 Animal cells

Abstract

The present invention relates to a method and apparatus of sorting objects including, providing a sample having wanted objects and unwanted objects; coating a surface of a sample holder with an antibody; placing an eluted sample on the sample holder; binding an antigen in the wanted objects with the antibody on the surface of the sample holder to sort the objects into wanted objects and unwanted objects; separating the wanted objects; and performing PCR-based STR analysis on the wanted objects. In one embodiment, holographic optical trapping is used to further sort the wanted objects. In other embodiments, the wanted objects are sperm and the antibody is a human sperm specific antibody, and the PCR is single cell PCR-based STR analysis. In still other embodiments, the binding is direct or indirect, ligands are used to bind to object-specific organomolecules, and protein A or protein G are used to bind the antibody.

Claims

1. A method of sorting objects comprising:

providing a sample having wanted objects and unwanted objects;

coating a surface of a sample holder with an antibody;

eluting said sample and placing said eluted sample on said sample holder;

binding an antigen in wanted objects in said sample with said antibody on said surface of said sample holder to sort said objects into bound wanted objects and unbound unwanted objects;

separating said wanted objects from said unwanted objects by removing said unwanted objects from a surface of said sample holder, leaving behind antibody-bound wanted objects on said surface of said sample holder;

inspecting and identifying said wanted objects, and determining a level of purity of said wanted objects;

releasing said wanted objects from said surface of said sample holder;

removing said wanted objects from said surface of said same sample holder using holographic optical trapping; and

performing single cell PCR-based STR analysis on said wanted objects in individual output chambers on said same sample holder.

2. The method of claim 1, wherein said wanted objects are cells, including sperm cells, said antibody is a human sperm specific antibody, and said unwanted objects are contaminants.

3. The method of claim 1,

wherein said PCR is bulk PCR on multiple sorted objects of a same type;

and

wherein said PCR performing step is compatible with automation and multiplexing.

4. The method of claim 1, further comprising:

performing said identifying step using one of brightfield microscopy or fluorescent microscopy.

5. The method of claim 1, further comprising:

binding an antigen to a primary antibody; and

binding said primary antibody to a secondary antibody.

6. The method of claim 1, further comprising:

immobilizing ligands on said sample holder;

binding cell surface receptors on a surface of said wanted objects with said ligands.

7. The method of claim 1 , further comprising:

binding a ligand to object-specific organomolecules; and

capturing said ligand bound object using an antibody.

8. The method of claim 2, wherein said sample holder contains beads coated with said antibody to bind, one of directly or indirectly, to said wanted objects.

9. The method of claim 8, further comprising:

binding said antigen to a primary antibody; and

binding said primary antibody to a secondary antibody, wherein said bead is coated with one of Protein A or Protein G, and

wherein said bead recognizes and captures said wanted objects bound to said primary antibody.

10. The method of claim 1, further comprising:

coating one of Protein A or Protein G on a surface of said sample holder;

binding said antigen to a primary antibody; and

binding said primary antibody to a secondary antibody;

wherein said Protein A or Protein G recognizes and captures said wanted objects that are antibody labeled.

11. The method of claim 1, wherein said sorting step using antibody-based sorting in conjunction with said holographic optical trapping separates cells of different types based on antigenic display.

12. The method of claim 1, further comprising:

generating a report with respect to said level of purity of said wanted objects prior to said PCR-based STR analysis.

13. The method of claim 1, wherein said same sample holder is an optically transparent microfluidics chip.

14. The method of claim 1, wherein said single cell PCR-based STR analysis is performed on said unwanted objects.

15. The method of claim 1, wherein said separating step includes washing said wanted objects from said sample holder.

16. The method of claim 2, wherein said removing step includes lysing said wanted objects from said sample holder.

17. The method of claim 2, wherein said removing step includes one of utilization of cleaving agents, altering/exchanging buffer solution, or altering a salt concentration to affect an effective charge shielding or charge switching.

18. The method of claim 17, wherein said removed wanted objects one of settle by gravity in said sample holder, or are pelleted down by centrifugation.

19. The method of claim 1, wherein said sorting of said wanted objects and unwanted objects is carried out in one of open or closed containers.

20. The method of claim 1, wherein said sample holder is a chip, and said single cell PCR-based STR or other DNA analysis is carried out on-chip.

21. The method of claim 1, wherein said inspecting step is performed using holographic optical trapping.

22. The method of claim 12, further comprising:

generating a final STR report for the wanted objects analyzed; and

generating statistics on an STR profile.

23. The method of claim 22, further comprising:

matching said STR profile data with a database for human identification.

24. The method of claim 5, further comprising:

incubating said wanted objects in excess antibody;

washing off said wanted objects by centrifugal spin; and

recovering said antibody labeled wanted objects on said sample holder.

25. The method of claim 5, wherein said secondary antibody is used to capture said wanted objects.

26. The method of claim 1, wherein said sample holder is one of optically transparent glass or plastic.

27. The method of claim 13, wherein said microfluidics chip contains an input chamber and said individual output chambers;

and

wherein each of said wanted objects are sorted using said holographic optical trapping, into each of said individual output chambers.