Method using a nonlinear optical technique for detection of interactions involving a conformational change

• US 8,497,073 B2
• Filed: 09/30/2009
• Issued: 07/30/2013
• Est. Priority Date: 01/24/2002
• Status: Active Grant

• Current Assignee: BlueLight Therapeutics, Inc.
• Original Assignee: Biodesy, LLC
• Inventors: Salafsky, Joshua S.
• Primary Examiner(s): LAM, ANN Y
• Application Number:
  US12/571,342
• Publication Number:
  US 20100120164A1
• Time in Patent Office: 1,399 Days
• Field of Search: None
• US Class Current: 435/7.1
• CPC Class Codes: B82Y 30/00 Nanotechnology for materials or surface science, e.g. nanocomposites
• CPC Class Codes: C12Q 1/6816 characterised by the detection means C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/6825 Nucleic acid detection involving sensors
• CPC Class Codes: G01N 21/31 Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry G01N21/72 takes precedence
• CPC Class Codes: G01N 21/636 using an arrangement of pump beam and probe beam; using the measurement of optical non-linear properties; non-linear optics per se G02F1/35
• CPC Class Codes: G01N 2333/726 G protein coupled receptor, e.g. TSHR-thyrotropin-receptor, LH/hCG receptor, FSH
• CPC Class Codes: G01N 2458/00 Labels used in chemical analysis of biological material
• CPC Class Codes: G01N 2500/02 Screening involving studying the effect of compounds C on the interaction between interacting molecules A and B (e.g. A = enzyme and B = substrate for A, or A = receptor and B = ligand for the receptor)
• CPC Class Codes: G01N 2500/04 Screening involving studying the effect of compounds C directly on molecule A (e.g. C are potential ligands for a receptor A, or potential substrates for an enzyme A)
• CPC Class Codes: G01N 33/5308 for analytes not provided for elsewhere, e.g. nucleic acids, uric acid, worms, mites
• CPC Class Codes: G01N 33/54313 the carrier being characterised by its particulate form
• CPC Class Codes: G01N 33/54373 involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings
• CPC Class Codes: G01N 33/56966 Animal cells
• CPC Class Codes: G01N 33/583 with non-fluorescent dye label
• CPC Class Codes: G01N 33/587 Nanoparticles
• CPC Class Codes: G01N 33/6845 Methods of identifying protein-protein interactions in protein mixtures
• CPC Class Codes: G01N 33/74 involving hormones or other non-cytokine intercellular protein regulatory factors such as growth factors, including receptors to hormones and growth factors
• CPC Class Codes: Y10T 436/143333 Saccharide [e.g., DNA, etc.]

Abstract

A nonlinear optical technique, such as second or third harmonic or sum or difference frequency generation, is used to detect binding interactions, or the degree or extent of binding, that comprise conformational change. In one aspect of the present invention, the nonlinear optical technique detects a conformational change in a probe due to target binding. In another aspect of the invention, the nonlinear optical technique screens candidate probes by detecting a conformational change due to a probe-target interaction. In another aspect of the invention, the nonlinear optical technique screens candidate modulators of a probe-target interaction by detecting a conformational change in the presence of the modulator.

Claims

1. A method for screening one or more candidate binding partners for modulating the conformation of a test molecule, the method comprising:

(a) illuminating a sample with one or more light beams at one or more fundamental frequencies, said sample comprising said test molecule exposed to said one or more candidate binding partners, wherein a nonlinear-active label is attached to a part of said test molecule that undergoes a conformational change upon interaction with said one or more binding partners, wherein there is no more than one test molecule directly bound to said nonlinear-active label, and wherein said test molecule is nonlinear-active in the absence of an exogenous nonlinear-active label bound to the test molecules;

(b) measuring one or more physical properties of a second-harmonic or sum-frequency light beam emanating from said sample; and

(c) comparing a value of said one or more physical properties measured in step (b) to a value for said one or more physical properties measured in the absence of exposure of said test molecule to said one or more candidate binding partners, wherein a change in the value of said one or more physical properties measured in step (b) relative to the value for said one or more physical properties measured in the absence of exposure of said test molecule to said one or more candidate binding partners indicates that said one or more candidate binding partners modulate the conformation of said test molecule.

2. A method for screening one or more candidate binding partners for modulating the conformation of a test molecule, the method comprising:

(a) illuminating a sample with one or more light beams at one or more fundamental frequencies, said sample comprising said test molecule exposed to said one or more candidate binding partners, wherein a nonlinear-active label is attached to a part of said test molecule that undergoes a conformational change upon interaction with said one or more binding partners, and wherein there is no more than one test molecule directly bound to said nonlinear-active label;

(b) measuring one or more physical properties of a second-harmonic or sum-frequency light beam emanating from said sample; and

(c) comparing a value of said one or more physical properties measured in step (b) to a value for said one or more physical properties measured in the absence of exposure of said test molecule to said one or more candidate binding partners, wherein a change in the value of said one or more physical properties measured in step (b) relative to the value for said one or more physical properties measured in the absence of exposure of said test molecule to said one or more candidate binding partners indicates that said one or more candidate binding partners modulate the conformation of said test molecule;

wherein the sample in step (b) further comprises at least one decorator with nonlinear activity that specifically binds to a candidate binding partner.

3. A method for screening one or more candidate binding partners for modulating the conformation of a test molecule, the method comprising:

(a) illuminating a sample with one or more light beams at one or more fundamental frequencies, said sample comprising said test molecule exposed to said one or more candidate binding partners, wherein a nonlinear-active label is attached to a part of said test molecule that undergoes a conformational change upon interaction with said one or more binding partners, and wherein there is no more than one test molecule directly bound to said nonlinear-active label;

(b) measuring one or more physical properties of a second-harmonic or sum-frequency light beam emanating from said sample; and

(c) comparing a value of said one or more physical properties measured in step (b) to a value for said one or more physical properties measured in the absence of exposure of said test molecule to said one or more candidate binding partners, wherein a change in the value of said one or more physical properties measured in step (b) relative to the value for said one or more physical properties measured in the absence of exposure of said test molecule to said one or more candidate binding partners indicates that said one or more candidate binding partners modulate the conformation of said test molecule

wherein the sample in step (b) further comprises at least one decorator with nonlinear activity that specifically binds to a test molecule.

4. A method for screening one or more candidate binding partners for modulating the conformation of a test molecule, the method comprising:

(a) illuminating a sample with one or more light beams at one or more fundamental frequencies, said sample comprising said test molecule exposed to said one or more candidate binding partners, wherein a nonlinear-active label is attached to a part of said test molecule that undergoes a conformational change upon interaction with said one or more binding partners, and wherein there is no more than one test molecule directly bound to said nonlinear-active label;

(b) measuring one or more physical properties of a second-harmonic or sum-frequency light beam emanating from said sample; and

(c) comparing a value of said one or more physical properties measured in step (b) to a value for said one or more physical properties measured in the absence of exposure of said test molecule to said one or more candidate binding partners, wherein a change in the value of said one or more physical properties measured in step (b) relative to the value for said one or more physical properties measured in the absence of exposure of said test molecule to said one or more candidate binding partners indicates that said one or more candidate binding partners modulate the conformation of said test molecule

wherein the sample in step (b) further comprises at least one decorator with nonlinear activity that specifically binds to a complex comprising one or more candidate binding partners bound to a test molecule.

5. A method for screening one or more candidate binding partners for modulating the conformation of a test molecule comprising:

(a) illuminating a sample with one or more light beams at one or more fundamental frequencies, said sample comprising said test molecule exposed to said one or more candidate binding partners, wherein a nonlinear-active label is attached to a part of said test molecule that undergoes a conformational change upon interaction with said one or more binding partners, wherein there is no more than one test molecule directly bound to said nonlinear-active label, and wherein said test molecule is nonlinear-active in the absence of an exogenous nonlinear-active label bound to the test molecules;

(b) measuring one or more physical properties of a second-harmonic or sum-frequency light beam emanating from a sample in step (a);

(c) illuminating a sample in step (a) with one or more light beams at one or more fundamental frequencies, said sample further comprising a known modulator of said test molecule;

(d) measuring one or more physical properties of a second-harmonic or sum-frequency light beam emanating from said a sample in step (c); and

(e) comparing a value of said one or more physical properties measured in step (b) to a value for said one or more physical properties measured in step (d), wherein a change in the value of said one or more physical properties measured in step (b) relative to the value for said one or more physical properties measured in step (d) indicates that said one or more candidate binding partners modulate the conformation of said test molecule.

6. A method for screening one or more candidate binding partners for modulating the conformation of a test molecule comprising:

(a) illuminating a sample with one or more light beams at one or more fundamental frequencies, said sample comprising said test molecule exposed to said one or more candidate binding partners, wherein a nonlinear-active label is attached to a part of said test molecule that undergoes a conformational change upon interaction with said one or more binding partners, and wherein there is no more than one test molecule directly bound to said nonlinear-active label;

(b) measuring one or more physical properties of a second-harmonic or sum-frequency light beam emanating from a sample in step (a);

(c) illuminating a sample in step (a) with one or more light beams at one or more fundamental frequencies, said sample further comprising a known modulator of said test molecule;

(d) measuring one or more physical properties of a second-harmonic or sum-frequency light beam emanating from said a sample in step (c); and

(e) comparing a value of said one or more physical properties measured in step (b) to a value for said one or more physical properties measured in step (d), wherein a change in the value of said one or more physical properties measured in step (b) relative to the value for said one or more physical properties measured in step (d) indicates that said one or more candidate binding partners modulate the conformation of said test molecule

wherein the sample in step (b) and/or step (d) further comprises at least one decorator with nonlinear activity that specifically binds to a candidate binding partner.

7. A method for screening one or more candidate binding partners for modulating the conformation of a test molecule comprising:

(a) illuminating a sample with one or more light beams at one or more fundamental frequencies, said sample comprising said test molecule exposed to said one or more candidate binding partners, wherein a nonlinear-active label is attached to a part of said test molecule that undergoes a conformational change upon interaction with said one or more binding partners, and wherein there is no more than one test molecule directly bound to said nonlinear-active label;

(b) measuring one or more physical properties of a second-harmonic or sum-frequency light beam emanating from a sample in step (a);

(c) illuminating a sample in step (a) with one or more light beams at one or more fundamental frequencies, said sample further comprising a known modulator of said test molecule;

(d) measuring one or more physical properties of a second-harmonic or sum-frequency light beam emanating from said a sample in step (c); and

(e) comparing a value of said one or more physical properties measured in step (b) to a value for said one or more physical properties measured in step (d), wherein a change in the value of said one or more physical properties measured in step (b) relative to the value for said one or more physical properties measured in step (d) indicates that said one or more candidate binding partners modulate the conformation of said test molecule

wherein the sample in step (b) and/or step (d) further comprises at least one decorator with nonlinear activity that specifically binds to a test molecule.

8. A method for screening one or more candidate binding partners for modulating the conformation of a test molecule comprising:

(a) illuminating a sample with one or more light beams at one or more fundamental frequencies, said sample comprising said test molecule exposed to said one or more candidate binding partners, wherein a nonlinear-active label is attached to a part of said test molecule that undergoes a conformational change upon interaction with said one or more binding partners, and wherein there is no more than one test molecule directly bound to said nonlinear-active label;

(b) measuring one or more physical properties of a second-harmonic or sum-frequency light beam emanating from a sample in step (a);

(c) illuminating a sample in step (a) with one or more light beams at one or more fundamental frequencies, said sample further comprising a known modulator of said test molecule;

(d) measuring one or more physical properties of a second-harmonic or sum-frequency light beam emanating from said a sample in step (c); and

(e) comparing a value of said one or more physical properties measured in step (b) to a value for said one or more physical properties measured in step (d), wherein a change in the value of said one or more physical properties measured in step (b) relative to the value for said one or more physical properties measured in step (d) indicates that said one or more candidate binding partners modulate the conformation of said test molecule

wherein the sample in step (b) and/or step (d) further comprises at least one decorator with nonlinear activity that specifically binds to a complex comprising one or more candidate binding partners bound to a test molecule.