Single-cell label-free assay

• US 8,703,428 B2
• Filed: 05/28/2008
• Issued: 04/22/2014
• Est. Priority Date: 10/06/2007
• Status: Expired due to Fees

• Current Assignee: Corning Incorporated
• Original Assignee: Corning Incorporated
• Inventors: Fang, Ye, Ferrie, Ann M., Wu, Qi
• Primary Examiner(s): Whisenant, Ethan C
• Application Number:
  US12/128,267
• Publication Number:
  US 20090093013A1
• Time in Patent Office: 2,155 Days
• Field of Search: 435/7.1, 435/243, 435/325, 435/410, 436/501
• US Class Current: 435/7.1
• CPC Class Codes: G01N 21/211 Ellipsometry optical thickness measurement G01B11/06
• CPC Class Codes: G01N 21/553 and using surface plasmons fluorescence excitation G01N21/648; enhanced Raman G01N21/658
• CPC Class Codes: G01N 21/554 detecting the surface plasmon resonance of nanostructured metals, e.g. localised surface plasmon resonance
• CPC Class Codes: G01N 33/5008 for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics
• CPC Class Codes: G01N 33/54373 involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings

Abstract

The disclosure provides a system and method for characterizing a single live-cell response to a stimulus with a biosensor imaging system having cells immobilized on the biosensor at a resolution level of a single cell, as defined herein.

Claims

1. A method for characterizing the response of live-cells a to a stimulus, the method comprising:

contacting the live-cells immobilized on a resonance waveguide grating (RWG) biosensor with a stimulus for a selected cellular target;

interrogating and detecting the dynamic mass redistribution (DMR) of each individual cell of the stimulus contacted live-cells using a spatially resolved swept wavelength RWG biosensor imaging system, wherein the system uses a wavelength tunable light source to illuminate and interrogate the biosensor and a digital camera to detect and record the resonant wavelength image of the RWG biosensor with a pixelated resolution less than the average size of the individual live-cells; and

determining the effect of the stimulus on the cell-signaling of the cellular target in the different individual cells among the stimulus contacted live-cells.

2. The method of claim 1 wherein the live-cells comprise at least one of:

an isolated single and individual cell, or a single and individual cell within a cell cluster.

3. The method of claim 1 wherein the effect of the stimulus on the cell-signaling of the cellular target in the stimulus contacted live-cells comprises comparing the dynamic mass redistribution of the cell-signaling of the cellular target in the presence and in the absence of the stimulus.

4. The method of claim 1 wherein the biosensor imaging system'"'"'s pixelated resolution is from about 1 to about 50 pixels per individual live-cell.

5. The method of claim 1 wherein the biosensor imaging system'"'"'s pixelated resolution is from about 1 to 10 pixels per individual live-cell at least in the direction perpendicular to the propagation direction of resonant light within the biosensor.

6. The method of claim 1 wherein each individual cell immobilized on the biosensor'"'"'s surface is one of a cluster of many cells when the cells have a confluency of from about 0.5% to about 100%.

7. The method of claim 1 wherein each individual cell immobilized on the biosensor'"'"'s surface is one of a cluster of many cells when the cells have a confluency of from about 80% to about 100%.

8. The method of claim 1 wherein the dynamic mass redistribution signal comprises an optical signal that measures real time kinetics of a stimulus-induced cellular response as a function of time.

9. The method of claim 1 wherein the dynamic mass redistribution signal comprises an optical signal that measures an endpoint or multiple points of a stimulus-induced cellular response over time and throughout a stimulation event.

10. The method of claim 1 wherein the biosensor imaging system provides biosensor output comprising at least one of:

the overall dynamics, the phase, the amplitude and kinetics of the phase, the transition time from one phase to another of the dynamic mass redistribution signal, or a combination thereof.

11. The method of claim 1 wherein the dynamic mass redistribution is the resonant wavelength or the resonant angle of the coupled light at each pixel or location of the biosensor and which wavelength or angle depends on the image interrogation mode of the biosensor system.

12. The method of claim 1 wherein the cellular target comprises a feature on the cell'"'"'s surface comprising at least one of a G protein-coupled receptor (GPCR), an ion channel, a receptor tyrosine kinase, a cytokine receptor, an immuno-receptor, an integrin receptor, an ion transporter, a pathogen recognition target, or a combination thereof.

13. The method of claim 1 wherein each individual cell immobilized on the biosensor'"'"'s surface is patterned onto the surface of the biosensor.

14. The method of claim 1 wherein the interrogating and detecting the dynamic mass redistribution (DMR) is accomplished in a period of time of at least one of:

from about seconds to about minutes, from about minutes to about hours, from about days to about weeks, or a combination thereof.

15. The method of claim 1 wherein the cellular target comprises an intracellular target comprising at least one of:

an enzyme, a kinase, a phosphatase, a monomeric or a dimeric receptor, an homologous or an heterologous receptor complex, or combinations thereof.

16. The method of claim 1 wherein each individual cell immobilized on the biosensor comprises an adherent cell, a suspension cell, or a combination thereof.

17. The method of claim 1 wherein the effect of the stimulus on the cell-signaling of the cellular target comprises comparing the stimulus-induced dynamic mass redistribution (DMR) of the cell-signalling of the cellular target in different individual cells to determine the homogeneity or heterogeneity of cellular responses at the single and individual cell level when the biosensor surface has more than one live-cell.

18. The method of claim 17 wherein the different individual cells are a single type of cells.

19. The method of claim 17 wherein the different individual cells contain at least two different types of cells.