SYSTEMS AND METHODS FOR HIGH THROUGHPUT ANALYSIS OF CONFORMATION IN BIOLOGICAL ENTITIES

• US 20150377900A1
• Filed: 06/29/2015
• Published: 12/31/2015
• Est. Priority Date: 06/30/2014
• Status: Active Grant

• Current Assignee: Quanta Therapeutics, Inc.
• Original Assignee: Biodesy, Inc.
• Inventors: Salafsky, Joshua, Dietz, Louis J., Bahatt, Dar, Hebert, Ray
• Granted Patent: US 9,989,534 B2
• Time in Patent Office: Days
• US Class Current: 1/1
• CPC Class Codes: B01L 2300/0636 Integrated biosensor, microarrays
• CPC Class Codes: B01L 2300/0819 Microarrays; Biochips
• CPC Class Codes: B01L 3/502715 characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
• CPC Class Codes: B01L 3/5085 for multiple samples, e.g. microtitration plates
• CPC Class Codes: G01N 21/552 Attenuated total reflection
• 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 33/54366 Apparatus specially adapted for solid-phase testing
• CPC Class Codes: G01N 33/54373 involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings
• CPC Class Codes: G01N 33/6845 Methods of identifying protein-protein interactions in protein mixtures
• CPC Class Codes: G02B 5/04 Prisms
• CPC Class Codes: G02B 5/045 Prism arrays

Abstract

Methods, devices, and systems are disclosed for performing high throughput analysis of conformational change in biological molecules or other biological entities using surface-selective nonlinear optical detection techniques.

Claims

1. A method for determining interactions between biological entities and test entities, comprising:

a) contacting each of at least four biological entities with at least one test entity;

b) illuminating each of the at least four biological entities with one or more excitation light beams using a surface selective optical technique; and

c) determining whether or not a conformational change was induced in each of the at least four biological entities through contact with the at least one test entity;

wherein determinations of conformational change are performed at an average rate of at least 10 test entities tested per hour.

2. The method of claim 1, wherein the determinations of conformational change are performed at an average rate of at least 100 test entities tested per hour.

3. The method of claim 1, wherein the at least four biological entities are the same.

4. The method of claim 1, wherein the at least four biological entities are different.

5. The method of claim 1, wherein the biological entities are selected from the group consisting of cells, proteins, peptides, receptors, enzymes, antibodies, DNA, RNA, oligonucleotides, small molecules, and carbohydrates, or any combination thereof.

6. The method of claim 1, wherein the biological entities are drug targets or portions thereof.

7. The method of claim 1, wherein each of the at least four biological entities are situated in a different discrete region on a substrate.

8. The method of claim 7, wherein each discrete region comprises an area of up to about 100 mm² on a substrate surface.

9. The method of claim 7, wherein each discrete region comprises a supported lipid bilayer.

10. The method of claim 9, wherein the biological entities are tethered to or embedded within the supported lipid bilayer.

11. The method of claim 1, wherein the at least one test entity is selected from the group consisting of cells, proteins, peptides, receptors, enzymes, antibodies, DNA, RNA, oligonucleotides, small molecules, and carbohydrates, or any combination thereof.

12. The method of claim 1, wherein the test entity is a drug candidate or portion thereof.

13. The method of claim 1, wherein the contacting step occurs in solution and comprises utilizing a pre-programmed fluid dispensing unit to dispense the at least one test entity.

14. The method of claim 1, wherein the contacting step comprises contacting each of the at least four biological entities with a different test entity.

15. The method of claim 1, wherein the contacting step comprises serially contacting each of the at least four biological entities with at least 100 different test entities.

16. The method of claim 1, wherein the contacting step comprises serially contacting each of the at least four biological entities with at least 10,000 different test entities.

17. The method of claim 1, wherein the one or more excitation light beams are provided by one or more lasers.

18. The method of claim 1, wherein the surface selective optical detection technique comprises the use of total internal reflection of at least one excitation light beam from a planar substrate surface.

19. The method of claim 1, wherein the determining step further comprises analyzing a non-linear optical signal.

20. The method of claim 19, wherein the non-linear optical signal is selected from the group consisting of second harmonic light, light, and difference frequency light.

21. The method of claim 7, further comprising moving said substrate relative to the position of one or more external sources of the one or more excitation light beams.

22. The method of claim 21, wherein each discrete region is optically coupled with an entrance prism and a different exit prism that are both integrated with a bottom surface of the substrate.

23. The method of claim 22, wherein the illuminating step comprises directing incident excitation light onto an entrance prism positioned adjacent to, but not directly below, each of the discrete regions.

24. The method of claim 23, further comprising collecting non-linear optical signals generated at each of the discrete regions upon illumination using an exit prism positioned adjacent to, but not directly below, each of the discrete regions, and directing said non-linear optical signals to a detector.

25. The method of claim 1, further comprising repeating the illuminating and determining steps a plurality of times after said contacting step, thereby determining conformational changes in each of the at least four biological entities as a function of time.

26. A device comprising:

a) a substrate, the substrate comprising;

i) an M×

N array of discrete regions formed on a surface of the substrate, wherein M is the number of rows of discrete regions and N is the number of columns of discrete regions in the array, and each discrete region is configured for containing a biological entity, and

ii) an R×

S array of prisms integrated with the substrate and optically coupled to the discrete regions, wherein R is the number of rows of prisms and S is the number of columns of prisms in the array;

wherein R=M+2 and S=N, or R=M and S=N+2.

27. The device of claim 26, wherein each of the discrete regions is optically coupled with at least one input prism and at least one output prism, and wherein the input prism and the output prism are spatially distinct.

28. The device of claim 26, wherein M=8 and N=12.

29. The device of claim 26, wherein M=16 and N=24.

30. The device of claim 26, wherein M=32 and N=48.

31. The device of claim 26, wherein M is greater than 4 and N is greater than 4.

32. The device of claim 26, wherein each discrete region comprises a supported lipid bilayer or is configured to facilitate the formation of a supported lipid bilayer.

33. The device of claim 26, further comprising a well-forming component bonded to a top surface of the substrate in order to isolate each discrete region in a separate well.

34. The device of claim 26, wherein each of the discrete regions comprises an area of up to about 100 mm².

35. The device of claim 26 or 33, wherein each discrete region or well is located directly above a single prism of the array of prisms integrated with the substrate.

36. The device of claim 26, wherein the substrate is composed of glass, fused-silica, or plastic.

37. An injection molding process for fabricating a prism array part from a plastic, the process comprising the use of two or more mold ejection devices to apply uniform pressure to the prism array part during a mold release step.

38. The injection molding process of claim 37, wherein the plastic is selected from the group consisting of cyclic olefin copolymer (COC), cyclic olefin polymer (COP), and acrylic.

39. The injection molding process of claim 37, wherein the two or more mold ejection devices impact the prism array part only in regions where the optical performance of the part is non-critical.

40. The injection molding process of claim 37, wherein the two or more mold ejection devices comprise an array of m×

n blade-like ejector features.

41. The injection molding process of claim 40, wherein m is greater than 2 and less than 20.

42. The injection molding process of claim 40, wherein n is greater than 2 and less than 20.