OPTICAL DETERMINATION OF LIVING VS. NON LIVING CELLS

• US 20100028853A1
• Filed: 11/02/2006
• Published: 02/04/2010
• Est. Priority Date: 10/25/2005
• Status: Abandoned Application

• Current Assignee: Board of Regents of Oklahoma State University
• Original Assignee: Board of Regents of Oklahoma State University
• Inventors: HARMON, HAROLD JAMES
• Application Number:
  US11/555,886
• Publication Number:
  US 20100028853A1
• Time in Patent Office: Days
• US Class Current: 435/5
• CPC Class Codes: G01N 33/5005 involving human or animal cells immunoassay G01N33/56966; immunoassays of protozoa G01N33/56905; protozoa in screening assays C12Q1/025
• CPC Class Codes: G01N 33/56911 Bacteria
• CPC Class Codes: G01N 33/56983 Viruses
• CPC Class Codes: G01N 33/583 with non-fluorescent dye label
• CPC Class Codes: G01N 33/84 involving inorganic compounds or pH

Abstract

A method of determining whether a cell sample in a medium contains living or dead cells of a predetermined target cell type is disclosed. The method includes preparing a testing substrate by attaching a binding molecule thereto, the binding molecule having the property of immobilizing cells of the target cell type upon coming in contact therewith. The method also includes incorporating a colorimetric indicator onto the testing substrate, performing a first spectrographic analysis of the calorimetric indicator, determining a change in pH of the medium based upon a second spectrographic analysis of the colorimetric indicator as compared to the first, and determining the portion of live target cells in the medium based upon the change in pH.

Claims

1. A method of determining whether a cell sample contains living or dead cells of a predetermined target cell type, the method comprising:

preparing a testing substrate by attaching a binding molecule thereto, the binding molecule having the property of immobilizing cells of the target cell type upon coming in contact therewith;

incorporating a calorimetric indicator onto the testing substrate;

performing a first spectrographic analysis of the colorimetric indicator;

determining a change in pH of the medium based upon a second spectrographic analysis of the colorimetric indicator as compared to the first; and

determining the portion of live target cells in the medium based upon the change in pH.

2. The method of claim 1, wherein the colorimetric indicator is a porphyrin.

3. The method of claim 1, wherein the calorimetric is a phthalocyanine.

4. The method of claim 1, wherein the colorimetric is a dye whose color changes with changes in pH

5. The method of claim 1, further comprising incorporating a second calorimetric indicator into the binding molecule and performing spectral analysis of the second calorimetric indicator to determine whether a cell of the target cell type has been bound on the testing substrate.

6. The method of claim 1, wherein the binding molecule is an antibody.

7. The method of claim 1, wherein the binding molecule is a natural receptor for the target cell type.

8. A method of determining whether a biological sample contains intact or fractured biologicals, the method comprising:

preparing a binding molecule, the binding molecule having the property of binding to a crytotope of the biological when exposed thereto;

incorporating a calorimetric indicator with the binding molecule to create a binding calorimetric hybrid, the colorimetric indicator having the property of displaying an altered spectrum when the cryptotope is bound with the binding molecule;

exposing the binding calorimetric hybrid to the biological sample; and

performing a spectral interrogation of the calorimetric indicator to determine whether the binding calorimetric hybrid has bound the cyptotope of the biological indicating a broken biological.

9. The method of claim 8, wherein the biological is a virus.

10. The method of claim 8, wherein the biological is a spore.

11. The method of claim 8, wherein the binding molecule is an antibody.

12. The method of claim 8, wherein the binding molecule is a natural receptor for the biological.

13. The method of claim 8, wherein the binding molecule is a substrate for an enzyme found an interior of the biological.

14. The method of claim 8, wherein the binding molecule is a substrate for an enzyme found on an exterior of the biological.

15. The method of claim 8, wherein the colorimetric indicator is selected from the group consisting of a porphyrin, a phthalocyanine, and a protein.

16. A method of determining if a medium contains intact or non-intact cells, the method comprising:

exposing a biological material sample to a first calorimetric indicator attached to a first molecule that can bind to a component on an exterior of a target cell;

exposing the biological material sample to a second calorimetric indicator attached to a second molecule that can bind to a component on an interior of the target cell;

interrogating the first calorimetric indicator over time to determine a number of exterior sites exposed that are bound to the first molecule;

interrogating the second calorimetric indicator over time to determine a number of interior sites exposed that are bound to the second molecule; and

determining the ratio of externally exposed components to internally located components.

17. The method of claim 16, wherein the interrogation is via fluorescence spectroscopy.

18. The method of claim 16, wherein the interrogation is via fluorescence microscopy.

19. The method of claim 16, wherein the interrogation is via absorbance spectroscopy.

20. The method of claim 16, wherein the first colorimeteric indicator is selected from the group consisting of a porphyrin, a phthalocyanine, and a protein.

21. The method of claim 16, wherein the second calorimetric indicator is selected from the group consisting of a porphyrin, a phthalocyanine, and a protein.

22. The method of claim 16, wherein the first colorimetric indicator attached to a first molecule that can bind to a component on the exterior of the cell is covalently bound to the molecule that can bind to a component on the exterior of the cell.

23. The method of claim 16, wherein the first molecule that can bind to a component on an exterior of a target cell is an antibody.

24. The method of claim 16, wherein the wherein the first molecule that can bind to a component on an exterior of a target cell is a natural receptor for the target cell.