Live-cell signals of pathogen intrusion and methods thereof

• US 20100323902A1
• Filed: 04/04/2008
• Published: 12/23/2010
• Est. Priority Date: 04/19/2007
• Status: Abandoned Application

• Current Assignee: Corning Incorporated
• Original Assignee: Corning Incorporated
• Inventors: Verrier, Florence, Lahiri, Joydeep, Fang, Ye
• Application Number:
  US12/080,765
• Publication Number:
  US 20100323902A1
• Time in Patent Office: Days
• US Class Current: 506/7
• CPC Class Codes: G01N 21/553 and using surface plasmons fluorescence excitation G01N21/648; enhanced Raman G01N21/658
• CPC Class Codes: G01N 33/54373 involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings
• CPC Class Codes: G01N 33/569 for microorganisms, e.g. protozoa, bacteria, viruses

Abstract

Disclosed is a system and method for measuring aspects of pathogen intrusion on a live-cell as defined herein. The system and method also provide a method to measure prophylaxis or remedial aspects of a therapeutic candidates in a live-cell or a live-cell model from pathogen intrusion.

Claims

1. A label-free method to detect pathogen intrusion in a live-cell, the method comprising:

providing an biosensor having a live-cell immobilized on a surface of the biosensor;

contacting the immobilized cell on the surface of the biosensor with a pathogen;

detecting a cell-signal pathway perturbation in a panel of markers that modulate distinct cellular targets; and

equating the extent of the perturbation with the extent of pathogen intrusion.

2. The method of claim 1 wherein the biosensor having a signal recognition element and a transducer element, comprises an evanescent wave device, an SPR device, an ellipsometric device, a reflectometric device, an electric impedance device, or combinations thereof.

3. The method of claim 1 the pathogen comprises at least one of a virus, a bacteria, a prion, or combinations thereof.

4. The method of claim 1 wherein the cell comprises a cell line, or a cell system.

5. The method of claim 1 wherein the pathogen intrusion comprises a cell'"'"'s response to at least one of a virus, a bacterium, a prion, or combinations thereof.

6. The method of claim 1 wherein the pathogen intrusion comprises an immune response of the cell to at least one of a virus, a bacterium, a prion, or combinations thereof.

7. The method of claim 1 wherein cell-signal pathway comprises at least one of a Ca²⁺ pathway, a mitogen-activated protein kinase pathway, an adhesion pathway, a cAMP pathway, an apoptotic pathway, cell cycle pathway, or combinations thereof.

8. The method of claim 1 wherein a marker comprises a molecule, a biomolecule, or a biological that can modulate an activity of at least one cellular target, and result in a reliably detectable biosensor output as measured by the biosensor.

9. The method of claim 8 wherein, when an evanescent wave biosensor is used, the biosensor output comprises a shift in resonant wavelength, a shift in resonant angle, or a change in peak width at half-maximum of the resonant peak.

10. The method of claim 8 wherein, when an electrical biosensor is used, the biosensor output comprises a change in bio-impedance.

11. The method of claim 8 wherein the cellular target comprises a receptor selected from the group consisting of a G_q-coupled receptor, a G_s-coupled receptor, a G_i-coupled receptor, a G_12/13-coupled receptor, an ion channel, a receptor tyrosine kinase, a transporter, a sodium-proton exchanger, a nuclear receptor, a cellular kinase, a cellular protein, and combinations thereof.

12. The method of claim 1 wherein the panel of markers comprises at least two markers and each marker modulates a cellular target selected from the group consisting of G_q-coupled receptor, a G_s-coupled receptor, a G_i-coupled receptor, a G_12/13-coupled receptor, an ion channel, a receptor tyrosine kinase, a transporter, a sodium-proton exchanger, a nuclear receptor, a cellular kinase, a cellular protein, and combinations thereof.

13. The method of claim 1 wherein the perturbation is a measure of, in a responsive cell:

the extent of pathogen intrusion;

the alteration in cellular activity attributable to pathogen intrusion;

the cell'"'"'s inflammatory response;

or combinations thereof.

14. The method of claim 1 further comprising contacting the immobilized cell on the surface of the biosensor with a prophylactic candidate or therapeutic candidate either before or after contacting the immobilized cell on the surface of the biosensor with a pathogen.

15. A method for characterizing the effect of a pathogen on a cell, the method comprising:

mapping a cell-signal network profile resulting from exposure of an immobilized cell to a pathogen according to claim 1;

comparing the mapped profile with a library of pathogen profiles; and

identifying a profile from the library of pathogen profiles that corresponds to the mapped profile.

16. The method of claim 15 wherein characterizing the effect of a pathogen comprises identifying a pathogen responsible for the effect.

17. The method of claim 15 wherein identifying a profile from the library of pathogen profiles comprises selecting a library profile that is an exact match or a best match of the mapped profile.

18. The method of claim 15 further comprising the step of contacting the immobilized cell with a prophylactic candidate or remedial candidate before or after the step of mapping the cell signaling network profile resulting from exposure of an immobilized cell to a pathogen.

20. The method of claim 18 wherein the pathogen comprises at least one of a virus, a bacterium, a prion, or combinations thereof.

19. A label-free method to detect a pathogen intrusion in a live-cell, the method comprising:

providing an optical biosensor having a live-cell immobilized on a surface of the optical biosensor;

contacting the immobilized cell on the surface of the biosensor with a pathogen; and

detecting a change in the cell'"'"'s local mass or local mass density within the detection zone of the biosensor relative to the cell prior to pathogen contact.

21. A method to monitor the effect of pathogen intrusion in a live-cell, the method comprising:

providing a live-cell having a pathogen intrusion to a biosensor surface;

culturing the live-cell having the pathogen intrusion with the biosensor surface until a defined confluency is achieved; and

measuring the biosensor output during the cell culture and intrusion.

22. A method to monitor the effect of pathogen intrusion in a live-cell, the method comprising:

providing a live-cell having a pathogen intrusion to a biosensor surface;

culturing the live-cell having the pathogen intrusion with the biosensor surface until a defined confluency is achieved; and

measuring the biosensor output for a predetermined and selected panel of markers.

23. The method of claim 22 wherein the biosensor continuously monitors at least one of the course of the pathogen intrusion, the marker-induced cell-signal changes, or both.