Measuring cellular metabolism of immobilized cells

• US 7,141,369 B2
• Filed: 04/25/2002
• Issued: 11/28/2006
• Est. Priority Date: 04/25/2002
• Status: Expired due to Term

• Current Assignee: Semibio Holdings Limited
• Original Assignee: Semibio Technology, Inc.
• Inventors: Cao, Bo
• Primary Examiner(s): FORMAN, BETTY J
• Application Number:
  US10/132,543
• Publication Number:
  US 20030203365A1
• Time in Patent Office: 1,678 Days
• Field of Search: 435/6, 435/7.1, 435/174, 435/283.1, 422/68.1, 530/300, 530/387.1
• US Class Current: 435/29
• CPC Class Codes: G01N 33/54353 with ligand attached to the carrier via a chemical coupling agent coatings G01N33/54393
• CPC Class Codes: G01N 33/56966 Animal cells
• CPC Class Codes: Y10T 428/315 Surface modified glass [e.g., tempered, strengthened, etc.]

Abstract

This invention uses a receptor molecule such as an antibody etc immobilized to a chip for a solid phase, and a sensor such as a pH electrode or an oxygen electrode or a glucose electrode for a search device. After immobilizing specific living cells by a bio-specific recognition reaction (a special immunity bond reaction) and washing the solid phase chip, one can measure the change of pH or oxygen consumption or glucose consumption from the metabolism of the specific living cells in substrate solution. This makes it possible to measure the activity and quantity of living cells specifically, accurately and quickly.

Claims

1. A method for obtaining at least one measurement of cellular metabolism of at least one specific living cell or at least one specific organelle in a sample, comprising the steps of:

a) providing a binder comprising;

(i) a planar flat-surfaced solid substrate comprised of quartz glass;

(ii) polyionic layers comprising an alternating arrangement of at least one polyanion layer and at least one polycation layer coated on and substantially parallel to the surface of said planar flat-surfaced solid substrate; and

(iii) at least one antibody or antibody fragment, affixed to the outermost of said polyionic layers and capable of binding a specific molecule located on said at least one specific living cell or said at least one specific organelle;

b) providing a sample suspected of containing said at least one specific living cell or said at least one specific organelle;

c) contacting said sample with said binder;

d) incubating said sample and said binder under conditions that allow said antibody or antibody fragment to bind said specific molecule, thereby immobilizing said at least one specific living cell or said at least one specific organelle;

e) providing a culture solution containing a substrate that can induce cellular metabolism;

f) contacting said culture solution with said immobilized at least one specific living cell or said immobilized at least one specific organelle; and

g) measuring change of pH of said culture solution to measure cellular metabolism.

2. The method of claim 1, wherein said planar flat-surfaced solid substrate is configured to comprises a capillary.

3. The method of claim 1, wherein said at least one antibody or antibody fragment is affixed to the outermost of said polyionic layers by covalent binding.

4. The method of claim 1, wherein said at least one antibody or antibody fragment is affixed to the outermost of said polyionic layers by non-covalent binding.

5. The method of claim 1, wherein said at least one antibody or antibody fragment is affixed to the outermost of said polyionic layers by biotin-avidin binding.

6. The method of claim 1, wherein said at least one antibody or antibody fragment is affixed to the outermost of said polyionic layers by aminosilane coupling.

7. The method of claim 1, wherein said sample comprises whole blood.

8. The method of claim 1, wherein said at least one specific living cell comprises a T-lymphocyte.

9. The method of claim 8, wherein said T-lymphocyte comprises a CD4⁺ T-lymphocyte.

10. The method of claim 1, wherein said specific molecule comprises a CD antigen.

11. The method of claim 10, wherein said CD antigen comprises a CD4 antigen.

12. The method of claim 1, further comprising pumping at least one solution selected from a sample solution, a wash solution, a reagent solution, a substrate solution, a labelling solution, and a combination thereof.

13. The method of claim 1, wherein the change of pH of said culture solution is measured with a pH sensitive field-effect transistor (pH-FET).

14. The method of claim 1, further comprising observing said immobilized at least one specific living cell or said immobilized at least one specific organelle under a microscope.