Method for Employing a Biosensor to Detect Small Molecules ("Fragments") that Bind Directly to Immobilized Protein Targets

• US 20080299673A1
• Filed: 04/19/2008
• Published: 12/04/2008
• Est. Priority Date: 04/19/2007
• Status: Abandoned Application

• Current Assignee: X-Body Incorporated (Bristol-Myers Squibb Company)
• Original Assignee: Sru Biosystems Incorporated
• Inventors: Wagner, Rick, Laing, Lance
• Application Number:
  US12/106,283
• Publication Number:
  US 20080299673A1
• Time in Patent Office: Days
• US Class Current: 436/171
• CPC Class Codes: G01N 2021/7773 Reflection
• CPC Class Codes: G01N 21/7743 the reagent-coated grating coupling light in or out of the waveguide
• CPC Class Codes: G01N 21/7746 the waveguide coupled to a cavity resonator
• CPC Class Codes: G01N 33/54373 involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings

Abstract

The invention provides method of detecting interactions of small molecules with target molecules.

Claims

1. A method of determining the affinity of small molecules for target molecules, wherein the method comprises:

(a) immobilizing the target molecules to a surface of a calorimetric resonant reflectance biosensor and determining a first peak wavelength value;

(b) adding the small molecules to the surface of the calorimetric resonant reflectance biosensor at 3 or more different concentrations and determining a peak wavelength values at the 3 or more different concentrations;

comparing the peak wavelengths to determine the affinity of the small molecules for the target molecules.

2. The method of claim 1, wherein the small molecules are less than about 300 Da.

3. The method of claim 1, wherein the dissociation constant (Kd) of the small molecules is determined.

4. The method of claim 3, wherein the dissociation constant of the small molecules is about 2,000, to about 750 μ

M.

5. The method of claim 1, wherein the concentration of the small molecules is about 0.5 mM to about 0.01 mM.

6. A method of determining a rank affinity of a small molecule sample comprising:

(a) detecting a first peak wavelength value of a surface of a colorimetric resonant reflectance biosensor;

(b) immobilizing target molecules with a known molecular weight to the surface of the colorimetric resonant reflectance biosensor and determining a second peak wavelength value;

(c) determining the amount of moles of the target molecule is bound to the calorimetric resonant reflectance biosensor using the first and second peak wavelength values;

(d) adding a specific concentration of a small molecule sample with a known molecular weight to the calorimetric resonant reflectance biosensor and determining a third peak wavelength value;

wherein the difference between the second peak wavelength value and the third peak wavelength value provides the amount of small molecule bound to the calorimetric resonant reflectance biosensor surface; and

(e) determining the rank affinity of the small molecule sample using ratio of the difference between the first peak wavelength value and the second peak wavelength value to the difference between the second peak wavelength value and the third peak wavelength value.

7. The method of claim 6, wherein the small molecule sample comprises small molecules that are less than about 300 Da.

8. The method of claim 6, wherein the dissociation constant of the small molecules in the small molecule sample is about 2,000, to about 750 μ

M.

9. The method of claim 6, wherein the concentration of the small molecules in the small molecule sample is about 0.5 mM to about 0.01 mM.

10. A method of determining if small molecules bind to a specific site on a target molecule or compete for target binding with a known blocker of a target site on the target molecule comprising:

(a) immobilizing the target molecule to a first calorimetric resonant reflectance biosensor;

(b) immobilizing the target molecule to a second calorimetric resonant reflectance biosensor, wherein the target site of the target molecule is blocked with a known blocker of the target site;

(c) adding small molecules to both the first and second colorimetric resonant reflectance biosensors and determining peak wavelength values for the first and second calorimetric resonant reflectance biosensors;

wherein, if the peak wavelength value of the first calorimetric resonant reflectance biosensor is shifted as compared to the peak wavelength value of the second calorimetric resonant reflectance biosensor, then the small molecules compete with the known blocker of a target site on the target molecule or binds to a specific site on the target molecule.

11. The method of claim 10, wherein the small molecules are less than about 300 Da.

12. The method of claim 10, wherein the dissociation constant of the small molecules is about 2,000, to about 750 μ

M.

13. The method of claim 10, wherein the concentration of the small molecules is about 0.5 mM to about 0.01 mM.

14. A method of determining if small molecules of less than 300 Da bind to target molecules, wherein the method comprises:

(a) immobilizing the target molecules to a surface of a calorimetric resonant reflectance biosensor and determining a first peak wavelength value;

(b) adding the small molecules to the surface of the calorimetric resonant reflectance biosensor and determining a second peak wavelength value;

comparing the first and second peak wavelengths, wherein if the second peak wavelength value is shifted as compared to the first peak wavelength value, then the small molecules bind the target molecules.

15. The method of claim 14, wherein the dissociation constant (Kd) of the small molecules is determined.

16. The method of claim 15, wherein the dissociation constant of the small molecules is about 2,000, to about 750 μ

M.

17. The method of claim 14, wherein the concentration of the small molecules is about 0.5 mM to about 0.01 mM.