Methods and Devices for Detection of Biological Materials Using Electric Field Assisted Rapid Analyte Capture

• US 20190317088A1
• Filed: 03/21/2019
• Published: 10/17/2019
• Est. Priority Date: 03/23/2018
• Status: Active Grant

• Current Assignee: University of Wyoming
• Original Assignee: University of Wyoming
• Inventors: DUTTA, Debashis
• Granted Patent: US 11,402,376 B2
• Time in Patent Office: Days
• CPC Class Codes: B01L 2300/0636 Integrated biosensor, microarrays
• CPC Class Codes: B01L 2300/0645 Electrodes
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• CPC Class Codes: B01L 2400/0418 electro-osmotic flow [EOF]
• CPC Class Codes: B01L 2400/0421 electrophoretic flow
• CPC Class Codes: B01L 3/502753 characterised by bulk separation arrangements on lab-on-a-chip devices, e.g. for filtration or centrifugation separation in general B01D; microapparatus for analysis using electrophoresis G01N27/44791; sample preparation G01N1/28
• CPC Class Codes: B01L 3/502776 specially adapted for focusing or laminating flows
• CPC Class Codes: G01N 21/69 specially adapted for fluids , e.g. molten metal
• CPC Class Codes: G01N 21/76 Chemiluminescence; Bioluminescence
• CPC Class Codes: G01N 33/54306 Solid-phase reaction mechanisms
• CPC Class Codes: G01N 33/54366 Apparatus specially adapted for solid-phase testing
• CPC Class Codes: G01N 33/5438 Electrodes
• CPC Class Codes: G01N 33/54393 Improving reaction conditions or stability, e.g. by coating or irradiation of surface, by reduction of non-specific binding, by promotion of specific binding

Abstract

In one embodiment, the present invention includes a system for detecting a target analyte which includes a microfluidic device having least one microfluidic channel with a binding surface positioned in the microfluidic channel with further include a first electrode and a second electrode. The system may further include a detector and a voltage supply. Also included is a method to detect a target analyte using a described microfluidics device, introducing solution with a target analyte to a binding surface, and binding the target analyte to the binding surface by applying an electrical potential between the first and second electrodes during at least a portion of the binding step, which enhances the rate of binding of the target analyte molecules to the binding molecules. The method then includes the steps of detecting a reporter molecule which corresponds to the amount of the bound target analyte molecules, which correlates with the amount of target analyte in the original sample. The method may also include multiple applications of sample to the binding surface prior to the detection step.

Claims

1. A system for detecting a target analyte, comprising a microfluidic device comprising:

(a) at least one microfluidic channel;

(b) a binding surface positioned in the microfluidic channel wherein at least a portion of the binding surface has channel walls comprising electrodes comprising a first electrode and a second electrode; and

(c) a detector positioned to detect a signal generated by the presence of the target analyte.

2. The system of claim 1, further comprising a voltage supply in electrical communication with the first and second electrodes in the at least one microfluidic channel.

3. The system of claim 1, further comprising a solution in the microfluidic device comprising a target analyte and a reporter molecule. Preliminary Amendment dated July 8, 2019

4. The system of claim 3, wherein the reporter molecule is a reporter antibody-catalyst conjugate capable of electrochemiluminescence reactions.

5. The system of claim 1, wherein the system further comprises:

(d) at least two microfluidic side channels positioned in fluid communication with the at least one microfluidic channels;

(e) a semipermeable membrane positioned in a first of the at least two of microfluidic side channels, the semipermeable membrane having a surface oriented towards the at last one microfluidic channel;

(f) a third electrode positioned in the first of the least two microfluidic side channels, and positioned such that the semipermeable membrane is positioned between the third electrode and the at least one microfluidic channel; and

(g) a fourth electrode positioned in the microfluidic channel or in a second of the at least two microfluidic side channels;

wherein (d), (e), (f) and (g) are positioned upstream of the detector.

6. The system of claim 5, wherein the semipermeable membrane comprises a silicate and a polymer.

7. A method to detect a target analyte in a sample, the method comprising the steps of:

(a) providing a microfluidic device comprising a binding surface in at least one microfluidic channel, wherein at least a portion of the binding surface has channel walls comprising electrodes comprising a first electrode and a second electrode, wherein the binding surface comprises binding molecules capable of selectively binding to the target analyte;

(b) introducing to the binding surface a first solution comprising the target analyte molecules, wherein at least a portion of the target analyte molecules can selectively bind to the binding molecules of the binding surface to provide a binding surface having bound target analyte molecules;

(c) binding the target analyte to the binding surface by applying an electrical potential between the first and second electrodes during at least a portion of the binding step, thereby enhancing the rate of binding of the target analyte molecules to the binding molecules;

(d) providing to the binding surface having the bound target analyte molecules a second solution comprising reporter molecules, wherein at least a portion of the reporter molecules further bind directly or indirectly to the target analyte molecules bound to the binding surface;

wherein the reporter molecules can generate a detectable signal; and

(e) detecting the signal indicating an amount of the originally present target analyte molecules in the sample.

9. The method of claim 7, wherein steps (b) and (c) are performed with additional solutions comprising the target analyte solution at least five times prior to performing step (d).

11. The method of claim 9, wherein at least 10 times more target analyte molecule is bound to the binding surface compared with binding the target analyte molecule in the absence of an applied electric field between the first and second electrodes.

14. The method of claim 7, wherein the amount of time required to bind a specified amount of target analyte molecules applied to the binding surface is reduced by approximately ten fold compared with binding the target analyte molecules in the absence of an applied electric field between the first and second electrodes.

17. The method of claim 7, wherein the binding surface is a derivatized glass with a silane compound and an aldehyde compound.

19. The method of claim 17, wherein the silane compound is 3-aminopropyltriethoxysilane and the aldehyde compound is glutaraldehyde.

20. The method of claim 7, wherein the binding molecules capable of selectively binding to the target analyte are an antibody with specificity for binding the target analyte.

21. The method of claim 20, wherein the method further comprises blocking nonspecific binding sites on the binding surface by applying a solution comprising a peptide, a polypeptide, or a combination thereof following step (a).

22. The method of claim 20, wherein the method further comprises removing unbound and nonspecifically bound target analyte following step (c).

23. The method of claim 20, wherein the method further comprises removing unbound and nonspecifically bound reporter molecules following step (d).

24. The method of claim 7, wherein the reporter molecules comprise an enzyme.

25. The method of claim 24, wherein the enzyme comprises alkaline phosphatase, acid phosphatase, horseradish peroxidase, beta-galactosidase, glucose oxidase, carboxypeptidase B, porcine liver esterase, rabbit esterase, lipase, butyryl cholinesterase, arginase, a catalyst for a bond cleavage reaction, a catalyst for a bond forming reaction, a catalyst for an oxidation reaction, a catalyst for a reduction reaction or any combination of these.

26. The method of claim 25, wherein the reporter molecules comprise a catalyst capable of electrochemiluminescence reactions.

27. The method of claim 26, wherein the catalyst is tris (bipyridine) ruthenium (II) chloride (Ru(bpy)3).

28. The method of claim 26, wherein the method further comprises applying an electrical potential between the first and second electrodes after the reporter molecule binding step (d) and wherein the detecting step (e) comprises detecting an electrochemiluminescence reaction.

29. The method of claim 24, wherein the reporter molecules comprise a catalyst capable of creating a colored or fluorescent ionized product from precursor colored or fluorescent product.

30. The method of claim 29, wherein the binding molecules on the binding surface comprise antigens of antibodies, antibodies, biotin, streptavidin, aptamers, nucleic acids, peptide nucleic acids, conjugates between antigens of antibodies, antibodies, biotin, streptavidin, aptamers, nucleic acids, peptide nucleic acids, or any combination of these.

31. The method of claim 7, wherein the method further comprises:

(e) providing to the binding surface having the bound target analyte molecules and the bound reporter molecules a solution comprising substrate molecules, wherein at least a portion of the substrate molecules undergo a catalytic chemical reaction initiated by the reporter molecules, thereby producing reaction product molecules having an ionic charge different from an ionic charge of the substrate molecules;

(f) transporting at least a some of the reaction product molecules into a microfluidic trapping region in fluid communication with the microfluidic binding region;

(g) concentrating the reaction product molecules in the microfluidic trapping region in front of, at the surface of and/or within a semipermeable membrane;

wherein the signal is measured from the reaction product molecules in front of, at the surface of and/or within a semipermeable membrane.

32. The method of claim 31, wherein the transporting step (f) comprises applying an electric potential between a third electrode positioned in a first microfluidic side channel and a fourth electrode positioned in the at least one microfluidic channel or in a second microfluidic side channel, wherein the semipermeable membrane is positioned in a first microfluidic side channel between the third electrode and the microfluidic channel and having a surface oriented towards the microfluidic channel;

and

wherein the electrical potential provides a force attracting the reaction product molecules towards the microfluidic trapping region.

33. The method of claim 31, wherein the step (g) is performed after one or more time intervals.

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