SMART GLASS SLIDE FOR MICROARRAYS

• US 20140274760A1
• Filed: 03/15/2013
• Published: 09/18/2014
• Est. Priority Date: 03/15/2013
• Status: Active Grant

• Current Assignee: Robert Bosch GmbH
• Original Assignee: Robert Bosch GmbH
• Inventors: KAVUSI, Sam, FOMINA, Nadezda, JOHNSON, Christopher, AHMAD, Habib
• Granted Patent: US 9,810,688 B2
• Time in Patent Office: Days
• US Class Current: 506/9
• CPC Class Codes: G01N 27/3272 Test elements therefor, i.e. disposable laminated substrates with electrodes, reagent and channels optical biosensors G01N33/52
• CPC Class Codes: G01N 33/5438 Electrodes

Abstract

Device for use in a biosensor comprising a multisite array of test sites, the device being useful for modulating the binding interactions between a (biomolecular) probe or detection agent and an analyte of interest from a biological by modulating the pH or ionic gradient near the electrodes in such biosensor. The device provides a biosensor which is more accurate, reliable and the results of which are more reproducible. Analytic methods for more accurately measuring an analyte of interest in a biological sample are also provided.

Claims

1. A device for use in a biosensor having a multisite array of test sites, the device comprising:

(a) a transparent support substrate supporting one or more electrodes; and

(b) a biomolecular interface layer having immobilized pH sensitive Fluorescent Protein and one or more immobilized probes thereon.

2. The device according to claim 1, wherein the pH sensitive Fluorescent Protein is a Green Fluorescent Protein (GFP).

3. The device according to claim 1, wherein the immobilized pH sensitive Fluorescent Protein covers both an area that is also covered by an electrode and an area that is not covered with an electrode.

4. The device according to claim 1, wherein the one or more electrodes is an indium tin oxide, gold, platinum, silver, or carbon electrode.

5. The device according to claim 1, comprising one or more counter-electrodes and wherein one or more electrodes are arranged in a multisite array, each site of the multisite array comprising a working electrode.

6. The device according to claim 5, wherein working electrode is an indium oxide electrode and the counter-electrode is selected from an indium oxide electrode, a gold electrode, a platinum electrode, a silver electrode, and a carbon electrode.

7. The device according to claim 1, wherein the immobilized probe is selected from a protein, a peptide, an antibody, an antigen, a nucleic acid, and a cell.

8. The device according to claim 7, wherein the one or more electrodes are arranged in a multisite array and each site comprises one or more immobilized probes.

9. The device according to claim 1, wherein the glass or plastic substrate placed in a biosensor is in contact with an aqueous solution comprising a phosphate buffer.

10. The device according to claim 9, wherein the aqueous solution comprises a diluted phosphate buffer and wherein the pH of the aqueous solution is between pH=5 to pH=8.

11. The device according to claim 9, wherein additional electrolytes selected from sodium sulfate, sodium or potassium chloride, sodium or potassium bromide, sodium or potassium iodide, sodium or potassium perchlorate, sodium or potassium nitrate, tetraalkylammonium bromide, tetraalkylammonium iodide, and combinations thereof are added to the aqueous solution.

12. The device according to claim 9, wherein the aqueous solution further comprises one or more buffer inhibitors.

13. The device according to claim 10, wherein the buffer inhibitor is a soluble polymer selected from poly(allylamine hydrochloride), poly(diallyldimethyl ammonium chloride), poly(vinylpyrroldone), poly(ethyleneimine), poly(vinylamine), poly(4-vinylpyridine) and tris(2-carboxyethyl)phosphine hydrochloride.

14. The device according to claim 9, wherein the aqueous solution comprises a water-miscible organic co-solvent selected from the groups consisting of acetonitrile, dimethyl sulfoxide (DMSO), dimethyl formamide (DMF), N,N-dimethyl acetamide (DMAc), and mixtures thereof.

15. The device according to claim 9, wherein the aqueous solution further comprises an electrochemically active agent selected from the group consisting of dopamine hydrochloride, ascorbic acid, phenol and derivatives thereof, benzoquinones and derivatives thereof, naphthoquinones and derivatives thereof, and 9,10-anthraquinone and derivatives thereof.

16. The device according to claim 1, wherein the one or more electrodes are connected to an electronic board via pogo-pins, a chip on foil via z-axis adhesive, or a chip on the substrate.

17. The device according to claim 16, wherein the electronic board or chip are powered by a printed battery, a small battery bound to the substrate, a magnetically coupled power transfer using coils on the substrate, or a rf-coupled power transfer using coils on the substrate.

18. The device according to claim 1, wherein the biosensor comprises a CMOS, electrode array, or TFT based system.

19. A biosensor comprising the device of claim 1.

20. The biosensor according to claim 19 comprising a multisite array of test sites in which the conditions for interacting with a biomolecule analyte can be varied independently at each test site, wherein the glass or plastic substrate is in contact with an aqueous solution comprising a phosphate buffer.

21. The biosensor according to claim 20, wherein additional electrolytes selected from sodium sulfate, sodium or potassium chloride, sodium or potassium bromide, sodium or potassium iodide, sodium or potassium perchlorate, sodium or potassium nitrate, tetraalkylammonium bromide, tetraalkylammonium iodide, and combinations thereof are added to the aqueous solution.

22. The biosensor according to claim 19, wherein the biosensor comprises a CMOS, electrode array, or TFT based system.

23. A method of modulating the pH or ionic concentration in a biosensor, the method comprising:

a) providing a biosensor according to claim 19 comprising a multisite array of test sites in which the conditions for interacting with a biomolecule analyte can be varied independently; and

b) reacting at the one or more electrodes an electrochemically active agent in an aqueous solution to produce H⁺ ion or OH⁻

ions.

24. The method of modulating the pH or ionic concentration in a biosensor according to claim 23, wherein the electrochemically active agent is added at a concentration of 1 nM to 100 mM.

25. The method of modulating the pH or ionic concentration in a biosensor according to claim 23, wherein the electrochemically active agent is selected from the group consisting of dopamine hydrochloride, ascorbic acid, phenol and derivatives thereof, benzoquinones and derivatives thereof, naphthoquinones and derivatives thereof, and 9,10-anthraquinone and derivatives thereof.

26. The method of modulating the pH or ionic concentration in a biosensor according to claim 23, wherein the aqueous solution comprises a dilute phosphate buffer and one or more additional electrolytes selected from sodium sulfate, sodium or potassium chloride, sodium or potassium bromide, sodium or potassium iodide, sodium or potassium perchlorate, sodium or potassium nitrate, tetraalkylammonium bromide and tetraalkylammonium iodide and optionally a buffer inhibitor.

27. The method of modulating the pH or ionic concentration in a biosensor according to claim 23, wherein the electrochemical reaction at the one or more electrode is carried out in a galvanostatic mode or potentiostatic mode.

28. The method of modulating the pH or ionic concentration in a biosensor according to claim 27, wherein the pH at the electrode is controlled through any one of an annealing pulse, pulse frequency, pulse width, and pulse shape.

29. The method of modulating the pH or ionic concentration in a biosensor according to claim 28, wherein the pulse width is in the range of 1 nanosecond to 60 minutes.

30. The method of modulating the pH or ionic concentration in a biosensor according to claim 23, wherein the pH close to the electrode surface is measured using the fluorescence intensity of surface-bound Green Fluorescent Protein (GFP).

32. The method for detecting a biomolecular analyte in a biological sample according to claim 30, wherein the Fluorescent Protein not located on or near an electrode is used as an internal reference for signal normalization.

31. A method for detecting a biomolecule analyte in a biological sample, the method comprising:

a) providing a biosensor comprising a multisite array of test sites in which the conditions for interacting with a biomolecule analyte can be varied independently, the biosensor having a device comprising a glass or plastic substrate supporting one or more electrodes and a biomolecular interface layer having immobilized pH sensitive Fluorescent Protein and one or more immobilized probes thereon, and at each test site having an aqueous solution comprising a dilute phosphate buffer and an electrochemically active agent;

b) at each test site electrochemically reacting the electrochemically active agent in an aqueous solution to produce H⁺ ion or OH⁻

ions, thereby modulating and controlling the pH at each test site;

c) adding a biological sample to each test site; and

d) detecting the biomolecule analyte in each test site,

wherein the amounts of electrochemically active agent and the electrochemical reaction are varied between test sites in a subset array of test sites in order to obtain sets of test sites in which the pH or ionic concentration near electrode surfaces in the test sites varies, and wherein the pH at each test site is determined by the fluorescence intensity of the pH sensitive Fluorescent Protein.