APPARATUS AND METHOD FOR QUANTIFYING BINDING AND DISSOCIATION KINETICS OF MOLECULAR INTERACTIONS

• US 20120295357A1
• Filed: 10/29/2010
• Published: 11/22/2012
• Est. Priority Date: 11/23/2009
• Status: Active Grant

• Current Assignee: Korea Research Institute of Standards and Science
• Original Assignee: Korea Research Institute of Standards and Science
• Inventors: Cho, Hyun Mo, Cho, Yong Jai, Che, Gal Won
• Granted Patent: US 8,940,538 B2
• Time in Patent Office: Days
• US Class Current: 436/34
• CPC Class Codes: B01L 2300/0654 Lenses; Optical fibres
• CPC Class Codes: B01L 2400/086 using baffles or other fixed flow obstructions
• CPC Class Codes: B01L 3/502715 characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
• CPC Class Codes: G01N 21/211 Ellipsometry optical thickness measurement G01B11/06
• CPC Class Codes: Y10T 436/143333 Saccharide [e.g., DNA, etc.]
• CPC Class Codes: Y10T 436/144444 Glucose

Abstract

The present invention relates to an apparatus for quantifying the binding and dissociation kinetics of molecular interactions of small molecular bio materials with high sensitivity almost without the influence of a change in the reflective index resulting from a buffer solution by making polarized incident light incident on the binding layer of a bio material, formed in a thin dielectric film, so that the polarized incident light satisfies a p-wave non-reflecting condition and a quantifying method using the same.

Claims

1. An apparatus for quantifying binding and dissociation kinetics of molecular interactions, comprising:

a micro flow path structure 100, comprising a support 110, a substrate 120 formed on the support 110 and made of a semiconductor or dielectric material, a thin dielectric film 130 formed on the substrate 120, a cover unit 140 configured to have an incident window 142 and a reflection window 144 respectively provided on one side and the other side and disposed on the support 110, and micro flow paths 150 formed in the support 110 and between the support 110 and the cover unit 140;

a sample injection unit 200 for forming a binding layer 160 of samples on the thin dielectric film 130 by injecting a buffer solution 210, comprising the samples of a bio material into the micro flow paths 150;

a polarization generation unit 300 for radiating incident light, polarized through the incident window 142, to the binding layer 160 at an angle of incidence θ

satisfying a p-wave non-reflecting condition; and

a polarization detection unit 400 for detecting a change in a polarization of reflected light of the binding layer 160 which is incident through the reflection window 144.

2. The apparatus according to claim 1, wherein the thin dielectric film 130 comprises a semiconductor oxide film or a glass film made of a transparent material.

3. The apparatus according to claim 2, wherein the thin dielectric film 130 has a thickness of 0 to 1000 nm.

4. The apparatus according to claim 1, wherein the micro flow path structure 100 comprises:

a plurality of inflow paths 152 and a plurality of outflow paths 154 formed on one side and the other side of the support 110, respectively, and

the micro flow paths 150 of multiples channels configured to have a plurality of barrier ribs 146 formed in an inner face of the cover unit 140 and to connect the inflow paths 152 and the outflow paths 154, respectively.

5. The apparatus according to claim 1, wherein:

the micro flow path structure 100 forms the micro flow path 150 of a single channel, and

a plurality of different self-assembled monolayers 132 to which the samples are bound is further formed on the thin dielectric film 130.

6. The apparatus according to claim 4, wherein:

each of the incident window 142 and the reflection window 144 of the cover unit 140 has a curved shell form having a predetermined curvature, and

the incident light and the reflected light are incident on the incident window 142 and the reflection window 144, respectively, at a vertical angle or at an almost vertical angle of the extent that a polarization state of each of the incident light and the reflected light is not greatly changed.

7. The apparatus according to claim 4, wherein:

each of the incident window 142 and the reflection window 144 of the cover unit 140 has a flat sheet shape, and

the incident light and the reflected light are incident on the incident window 142 and the reflection window 144, respectively, at a vertical angle or at an almost vertical angle of the extent that a polarization state of each of the incident light and the reflected light is not greatly changed.

8. The apparatus according to claim 4, wherein the cover unit 140 is integrally made of glass or a transparent synthetic resin material.

9. The apparatus according to claim 1, wherein the binding layer 160 is a multi-layered film, comprising self-assembled monolayers 132 suitable for bonding characteristics of various bio materials, a fixation material, and various bio materials including small molecules bonded to the fixation material.

10. The apparatus according to claim 1, wherein the polarization generation unit 300 comprises:

a light source 310 for radiating predetermined light, and

a polarizer 320 for polarizing the radiated light.

11. The apparatus according to claim 10, wherein the light source 310 radiates monochromatic light or white light.

12. The apparatus according to claim 10, wherein the light source 310 is a laser or a laser diode having a wavelength-variable structure.

13. The apparatus according to claim 10, wherein the polarization generation unit 300 comprises at least one of:

a collimation lens 330 for providing parallel light to the polarizer 320,

a focusing lens 340 for increasing an amount of the incident light by converging the parallel light passing through the polarizer 320, and

a first compensator 350 for phase-delaying polarized components of the incident light.

14. The apparatus according to claim 13, wherein the polarizer 320 and the first compensator 350 are rotatably configured or further equipped with other polarization modulation means.

15. The apparatus according to claim 1, wherein the polarization detection unit 400 comprises:

an analyzer 410 configured to polarize the reflected light,

a photodetector 420 configured to obtain predetermined optical data by detecting the polarized reflected light, and

an operation processor 430 electrically connected to the photodetector 420 and configured to induce quantified values based on the optical data.

16. The apparatus according to claim 15, wherein the photodetector 420 comprises any one of a CCD type solid state array, a photomultiplier tube, and a silicon photodiode.

17. The apparatus according to claim 15, wherein the operation processor 430 induces the quantified values, including a binding concentration, a binding constant, and a dissociation constant of the samples, by finding an ellipsometric constant Ψ

or Δ

18. The apparatus according to claim 15, wherein the polarization detection unit 400 further comprises at least one of:

a second compensator 440 for phase-delaying polarized components of the reflected light, and

a spectroscope 450 for making spectroscopic the reflected light.

19. The apparatus according to claim 18, wherein the analyzer 410 and the second compensator 440 are rotatably configured or further equipped with other polarization modulation means.

20. A method of quantifying binding and dissociation kinetics of molecular interactions, the method comprising:

a first step S100 of a sample injection unit 200 injecting a buffer solution 210, including samples of small molecular bio materials, into micro flow paths 150 of a micro flow path structure 100;

a second step S200 of the samples being bound to a thin dielectric film 130 of the micro flow path structure 100, thus forming a binding layer 160;

a third step S300 of a polarization generation unit 300 polarizing predetermined light and making the polarized light incident on the binding layer 160 at an angle of incidence to satisfy a p-wave non-reflecting condition through an incident window 142 of the micro flow path structure 100;

a fourth step S400 of reflected light of the binding layer 160 being incident on a polarization detection unit 400 through a reflection window 144 of the micro flow path structure 100; and

a fifth step S500 of the polarization detection unit 400 detecting a polarization state of the reflected light using an ellipsometry or a reflectometry.

21. The method according to claim 20, wherein in the first step S100, the buffer solution 210 including the samples of different concentrations is injected into respective micro flow paths 150 of the micro flow path structure 100 including multiple channels.

22. The method according to claim 20, wherein in the second step S200, the samples are bound to a plurality of different self-assembled monolayers 132 formed on the thin dielectric film 130, thus forming the different binding layers 160.

23. The method according to claim 20, wherein the fifth step S500 comprises:

a step of an analyzer 410 polarizing the reflected light,

a step of a photodetector 420 obtaining predetermined optical data by detecting the polarized reflected light, and

a step of an operation processor 430 inducing quantified values, including a binding concentration, a binding constant, and a dissociation constant of the samples, by finding an ellipsometric constant Ψ

or Δ

of the ellipsometry on the basis of the optical data.