Electrochemical Device

Abstract

An electrochemical microsensor comprising an array of working microelectrodes, the working microelectrodes include: one or more bare microelectrodes; one or more thick film-coated microelectrodes, optionally with conductive additive incorporated into the coating, selected from the group consisting of polysaccharide-coated microelectrodes and platinum black-coated microelectrodes; one or more thin film-coated microelectrodes selected from the group consisting of reduced graphene oxide-coated microelectrode and transition metal chalcogenide-coated microelectrodes; wherein the electrochemical microsensor further comprises a counter electrode and optionally one or more reference microelectrode(s).

Claims

1. An electrochemical microsensor comprising an array of working microelectrodes, the working microelectrodes include: one or more bare microelectrodes; one or more thick film-coated microelectrodes, optionally with conductive additive incorporated into the coating, selected from the group consisting of polysaccharide-coated microelectrodes and platinum black-coated microelectrodes; one or more thin film-coated microelectrodes selected from the group consisting of reduced graphene oxide-coated microelectrode and transition metal chalcogenide-coated microelectrodes; wherein the electrochemical microsensor further comprises a counter electrode and optionally one or more reference microelectrode(s).

2. An electrochemical microsensor according to claim 1, wherein the array of working microelectrodes comprises: one or more bare microelectrodes; one or more polysaccharide-coated microelectrodes; one or more polysaccharide-coated microelectrodes with conductive additives incorporated into the coating; one or more platinum black-coated microelectrodes; one or more reduced graphene oxide-coated microelectrodes; one or more MoS.sub.2-coated microelectrodes; and one or more WS.sub.2-coated microelectrodes.

3. An electrochemical microsensor according to claim 1 in the form of lab-on-a-chip.

4. An electrochemical microsensor according to claim 3, wherein the chip comprises a base substrate and has a recessed region on its surface, defined by a wall made of electrically insulating polymer which is elevated in respect of said recessed region, such that the recessed region can serve as a receptable for a liquid sample, with discrete microstructures placed inside said recessed region, wherein a microstructure consists of a microelectrode deposited atop of an adhesion layer attached to the base substrate, wherein each microstructure is encircled by a wall made of electrically insulating polymer, thereby defining a plurality of microchambers, the interior of which is occupied by the microstructures.

5. An electrochemical microsensor according to claim 4, wherein the counter electrode is located inside the recessed region.

6. An electrochemical microsensor according to claim 5, wherein the recessed region has a circular shape, with the counter electrode being positioned concentrically in the recessed region and the microstructures placed along the perimeter of the recessed region.

7. A device for electrochemical detection, comprising: an electrochemical microsensor as defined in claim 1; a potentiostat or galvanostat to which the working microelectrodes, the counter electrode and optionally the reference microelectrodes are electrically connected to allow control of the potential or current of the working microelectrodes, respectively, to create a data set of electrochemical signals when the microelectrodes are immersed in a sample; a processor configured to analyze the data set of electrochemical signals by one or more chemometric techniques.

8. A device according to claim 7, comprising a potentiostat that records current signals measured by voltammetry.

Description

[0098] In the drawings:

[0099] FIG. 1 shows the electrochemical detectability of clozapine in a blood sample using a single surface-modified electrode to generate an electrochemical signal indicative of clozapine concentration.

[0100] FIG. 2 shows the electrochemical detectability of clozapine in a blood sample using an array of multiple surface-modified electrodes to generate an electrochemical signal indicative of clozapine concentration according to the invention.

[0101] FIG. 3 shows a design of the multielectrode array patterned on a chip (lab-on-chip).

[0102] FIG. 4 schematically illustrates the fabrication of the multielectrode array using photolithography and metal deposition.

[0103] FIG. 5 is a photograph showing the electrical chip of the invention and a potentiostate.

[0104] FIG. 6 shows the modifications of gold microelectrodes in the multi-electrode array: (A) Ag (B) AgCl, (C) Pt-black, (D) r-GO, (E) and (F) MoS.sub.2, (G) and (H) WS.sub.2, (I) chitosan, and (J) chitosan-CNT. (K) Optical image of the multi-electrode array.

[0105] FIG. 7 shows the SEM characterization of the multi-electrode array. Low and high resolution image of the (A) and (B) bare, (C) and (D) Ag/AgCl, (E) and (F) Pt-black, (G) and (H) r-GO, (I) and (J) MoS.sub.2, (K) and (L) WS.sub.2, (M) and (N) chitosan, (O) and (P) chitosan-CNT.

[0106] FIG. 8 show electrochemical characterization of the multi-electrode array. (A) Linear regression of the OCP vs log(Cl.sup.−), CVs recorded in 10 mM PBS solution using (B) Pt-black -modified and (C) chitosan, chitosan-CNT, r-GO, MoS.sub.2, WS.sub.2 -modified and bare microelectrodes in the multielectrode array. (D) CVs recorded in 5 mM ferricyanide/ferrocyanide solution using multi-electrode array, (E) intra-chip reproducibility for the oxidation peak current for each modification, and (F) intra-chip reproducibility for the thickness measurement in the multielectrode array

[0107] FIG. 9 illustrates measuring clozapine in capillary whole blood samples of schizophrenia patients by the multielectrode array (DPV), using (A) Pt-black (blue), and (B) Chitosan (light black), chitosan-CNT (red), r-GO (light blue), MoS.sub.2 (green), WS.sub.2 (light purple) -modified and bare microelectrodes (dark yellow) in the multi-electrode array, (C) PRESS vs the factors, and (D) Linear regression of estimated vs the actual CLZ concentrations using the PLSR model.

[0108] FIG. 10 is a schematic illustration of the device of the invention suitable for use as point-of-care testing device.

EXAMPLES

Preparations 1-7

Electrodeposition Solutions

[0109] 1) Ag and HCl Solutions

[0110] Silver nitrate, Ammonium carbonate, 5-Sulfosalicylic Acid Dihydrate, and Piperazine were mixed in the ratio of 2:1:6:2 in DI water. The solution was stirred at 400 rpm for 10 minutes. The solution pH was adjusted to 9.5 by adding 0.25 M ammonium solution in several steps. The 0.25 M ammonium solution used to adjust the pH was prepared from ammonium hydroxide. 0.1 M HCl solution was prepared from 10.2 M HCl stock solution which was used to chlorinate the Ag to Ag/AgCl surface.

[0111] In addition, KCl solution was prepared to characterize the Ag/AgCl reference microelectrode. 1 M KCl stock solution was prepared from KCl powder, which was further diluted to 0.5, 0.1, and 0.001 M different concentrations.

[0112] 2) Platinum-Black Electrodeposition Solution

[0113] Platinum black deposition solution was prepared by mixing 0.5 g of chloroplatinic acid and 25 mg of lead acetate in 50 ml of DI water. The mixture was then stirred and 3.9 μL of concentrated hydrochloric acid (32%; 10.2 Molar concentration) was added to the mixture. The prepared solution was covered with aluminum foil and stored at room temperature.

[0114] 3) Graphene Oxide Electrodeposition Solution

[0115] Graphene oxide (GO) solution was prepared using a modified Hummers' method. A 9:1 ratio of sulfuric acid and phosphoric acid (100 mL) was prepared and stirred for several minutes. A graphite powder (7.5 g/L, 1 wt. eq.) was added to the mixture under stirring conditions. Potassium permanganate (45 g/L, 6 wt. eq.) was slowly added to the solution and the mixture was stirred for 6 h at 30-35° C. until the color turned to dark green. To eliminate the excess of potassium permanganate, hydrogen peroxide 30% w/w (2.5 mL) was added slowly and the mixture was stirred for 10 min, resulting in an exothermic reaction that was left to cool at room temperature. Concentrated 32% hydrochloric acid and DI were sequentially added at a 1:3 volume ratio and the resulting solution was centrifuged at 7000 RPM for 5 min. Residuals of the centrifuged solution were washed 3 times with hydrochloric acid and DI (1:3 v/v). The washed GO solution was dried at 90° C. in an oven (Binder-9010-0082) overnight, yielding the GO powder. Dried GO powder was dissolved in DI, resulting in an electrodeposition solution with 0.5 g/L GO concentration. Next, 100 mM sodium chloride solution was added as an electrolyte to the GO solution, resulting in a final GO electrodeposition solution. At higher GO concentrations, rapid precipitation was observed that prevented the efficient electrodeposition of r-GO.

[0116] 4) MoS.sub.2 Electrodeposition Solution

[0117] 0.1 mg/mL MoS.sub.2 solution was prepared from 1 mg/mL stock monolayer MoS.sub.2 solution in 0.1 M H.sub.2SO.sub.4 solution and DI water. The solution was sonicated for 10 minutes. 0.1M H.sub.2SO.sub.4 solution was prepared from 18.4 M stock H.sub.2SO.sub.4 solution.

[0118] 5) WS.sub.2 Electrodeposition Solution

[0119] 0.1 mg/mL WS.sub.2 solution was prepared from 1 mg/mL stock monolayer WS.sub.2 solution in 0.1 M H.sub.2SO.sub.4 solution and DI water. The solution was sonicated for 10 minutes before electrodeposition.

[0120] 6) Chitosan electrodeposition solution

[0121] A concentrated chitosan solution (1.8%, pH 5.5) was prepared by dissolving chitosan flakes in HCl (2 M) to achieve a final pH of 5-6. Then, the concentrated chitosan solution was diluted with Mili-Q water to obtain a chitosan solution (1%).

[0122] 7)Chitosan-Carbon Nanotube Electrodeposition Solution

[0123] Chitosan-CNT electrodeposition solution was prepared by mixing the chitosan solution (1%, 100 mL) with CNTs (1.75 g, 1.75%), followed by an ultra-sonication (ElmasonicS10H, Elma) step of 20 min and then stored at 4° C.

Example 1

Microfabrication of an Array of Surface-Modified Gold Microelectrodes on Glass Substrate

Step 1: Creating an Array of Bare Gold Microelectrodes

[0124] A borosilicate glass substrate was cleaned with acetone, isopropanol, and DI water and then dried with nitrogen gas (FIG. 4A(i)). The photoresist (AZ 5214E) coating process started with spinning the wafer with the photoresist at 2200 RPM for 12 s, followed by a soft bake on a contact hot plate at 110° C. for 2 min and 30 s. Next, the electrode patterns were transferred from the mask using a hard contact of 7.6 mW/cm.sup.2 for 65 s using a mask aligner (Karl Suss Mask Aligner MA6). The exposed wafer was then developed in AZ 726 MIF developer for 2 min, followed by rinsing in DI water for 5 min (FIG. 4A(ii-iii)). Next, 20 nm of titanium and 200 nm of gold were deposited using the E-gun deposition system (FIG. 4A(iv)). The wafer was then transferred to a beaker with acetone solution for a lift of process that resulted in the Au/Ti microelectrode patterns on a glass substrate (FIG. 4A(v)). The wafer was again rinsed in the DI water for 1 min to remove any residue from the wafer. SU8-3005 was used to define the microelectrode chamber; this allows cleaning the microelectrode with an AMI (acetone, methanol, and isopropanol) solution without destroying the chamber before using it. The process starts with spin coating SU8-3005 at 3000 RPM for 30 s, followed by a soft bake on a contact hot plate at 95° C. for 15 min. Next, the photoresist was exposed to light through the electrodes' mask using a hard contact of 7.6 mW/cm.sup.2 for 50 s at a Mask Aligner (MA6, SUSS MicroTec). Post Exposer Bake (PEB) for 5 min at 95° C. was used, since this is a negative photoresist and the wafer was cooled down to room temperature. (FIG. 4B (i)). The exposed wafer was then developed in PGMA ERB developer solution for 8 min and washed in isopropanol solution for 10 s. The hard bake on a contact hot plate at 150° C. for 5 min was carried out to remove any hydration on the substrate, and oxygen plasma cleaning (30 W, 500 mTorr, 2 min, 3 sccm) was used after the hard bake to remove any residues or impurities on the substrate (FIG. 4B (ii)). The process of chamber microfabrication was repeated with same parameter but with new SU8 3050 to define the big chamber for solution (FIG. 4B (iii and iv)). The wafer was then coated with photoresist before dicing it into chips using a Dicing saw (ADT-7100). All chips were cleaned with AMI solution before use. The optical image was recorded using optical microscope.

Step 2: Surface Modification of the Microelectrodes

[0125] VSP potentiostat (Bio-Logic, Ltd.) was used for the modification of the microelectrodes to create different coatings on the gold surfaces in the multi-electrode array. Coating were electrodeposited using three-electrode cell arrangement or two-electrode cell arrangement.

[0126] The three-electrode cell configuration consisted of the microfabricated gold microelectrode(s) as working electrode, an externally applied commercial Pt ring counter electrode with an approximate surface area of 3.6 cm.sup.2 (CHI115, CH Instruments; counter electrode; ‘CE’), and an Ag/AgCl 3 M NaCl reference electrode (CHI111, CH Instruments; reference electrode; ‘RE’ E.sub.SHE=0.210+E.sub.Ag/AgCl).

[0127] For the two-electrode configuration (chronopotentiometry) reference electrode was shorted to the Pt ring electrode. All electrochemical potential values are versus Ag/AgCl half-cell potential.

Ag/AgCl Coated Microelectrodes (n.SUB.Ag/AgCl.=2)

[0128] 40 mL of Ag solution (Preparation 1) was used under continuous stirring at 200 rpm to electroplate gold microelectrode surface with Ag, applying the chronopotentiometry technique (cathodic current of 1.5 μA for 10 minutes) to selectively electroplate two microelectrodes simultaneously, by the two-electrode configuration, i.e., with reference electrode connected to ring counter electrode and two working microelectrodes from the chip. After the Ag electroplating, ⅓ to ¼ of the Ag surface was converted to AgCl by chlorination in 0.1 M HCl solution. For this purpose, the total amount of charge, required to convert ⅓ to ¼ portion of electroplated Ag surface to Ag/Cl was calculated. A chronoamperometry technique was applied, at constant potential of 0.22 V under continuous stirring condition at 200 rpm for this purpose. Three electrode system, ring counter, Ag/AgCl reference and Ag electroplated as working microelectrodes were used (equation 1-4).

Q.sub.Ag=I.sub.appiled×Time, Q.sub.req_max=Q.sub.Ag (⅓) to Q.sub.req_min=Q.sub.Ag (¼);   (1)

Example: Q.sub.Ag=1.570 μA×600 Sec=942 μQ;   (2)

Q.sub.req_max=Q.sub.Ag(⅓)=942 μQ×(⅓)=314 μQ;   (3)

Q.sub.req_min=Q.sub.Ag(¼)=942 μQ×(¼)=235.5 μQ;   (4)

[0129] Hence, the amount of charge that needs to be transferred is between 235.5 and 314 μQ using chronoamperometry technique.

[0130] FIG. 6A shows the chronopotentiogram of the Ag electroplating onto the Au microelectrode for 10 min. This resulted in two Au surface electroplated with Ag. FIG. 6B shows the chlorination of the two Ag electroplated Au surfaces at a constant potential to Ag/AgCl, by converting the Ag surface to AgCl via charge transfer mechanism.

[0131] Open circuit potential (OCP; for 5 minutes) of the Ag/AgCl electroplated gold microelectrode was recorded in different concentration solution of KCl vs the commercial ref. electrode. The recorded potential was plotted against the logarithm of chloride concentration. The slope of the linear regression of the potential vs log[Cl.sup.−1] was compared with the standard Randle-Sevcik equation to calculate the slope and standard potential (FIG. 8A). The slope of the linear regression was found to be 0.045 V whereas the intercept was found to be 0.03 V. The results indicate that the so-formed Ag/AgCl electroplated gold microelectrode could serve as reference electrodes.

Platinum-Black Coated Microelectrodes (n.SUB.Pt.=3)

[0132] Before the gold microelectrodes were modified with the aid of the electrodeposition solution of Preparation 2, the chip was cleaned with AMI solution, followed by rinsing in DI water. The chip was then dried with nitrogen. The chronopotentiometry technique (cathodic current density=4.8 mA/cm.sup.2, time=5 min) was used to modify gold microelectrodes to create Pt coating. A constant cathodic current (equivalent to current density) of 7.5 μA was applied to three gold microelectrode simultaneously. This allows the deposition of Pt on the gold surface., i.e., by the electrostatic interaction between Pt ions in the solution and the negatively charged Au surface. The chronopotentiogram is shown in FIG. 6C.

Reduced Graphene Oxide Coated Microelectrodes (n.SUB.r-GO.=3)

[0133] 50 μL of the GO electrodeposition solution (Preparation 3) was dropped on the chamber and the r-GO was selectively deposited using cyclic voltammetry (CV) technique (scanning initial potential E.sub.i=−1.4 V vs RE, vertex #1 potential E.sub.1=−1.4 V vs RE, vertex #2 potential E.sub.2=1.4 V vs RE, scan rate=0.1 V/s, and the number of cycles=3). The electrodeposition process was optimized by varying the number of cycles (1, 2, 3, 4, and 5) and the scan rate (0.05, 0.1, 0.2, 0.3, and 0.5 V/s. Electrochemical modifications were performed using a VSP potentiostat (Bio-Logic, Ltd.) and in a three-electrode cell configuration consisting of the microfabricated microelectrode (working electrode; ‘WE’), an externally applied commercial Pt wire (CHI115, CH Instruments; counter electrode; ‘CE’), and a Tungsten needle (P/N H-20242, Quarter) coated with Ag/AgCl ink (011464, BAS Inc.; pseudo reference electrode; ‘RE’). All electrochemical potential values are versus Ag/AgCl half-cell potential. FIG. 6D shows the CVs scanned over −1.4V to 1.4V at rate of 100 mV/s for three cycles for the electrodeposition of r-GO film on three Au microelectrodes simultaneously.

MoS.SUB.2 .Coated Microelectrodes (n.SUB.MoS2.=3)

[0134] The gold multi-microelectrode array chip was dipped in a 10 mL solution of MoS.sub.2 (1 g/L) dispersed in 0.1 M sulfuric acid (see Preparation 4). The material was selectively deposited using CV technique. Prior to the electrodeposition, the gold microelectrodes were electrochemically cleaned using a CV in a 0.5 M H.sub.2SO.sub.4 electrolyte by cycling the potential from −0.4 to 1.4 V.sub.Ag/AgCl for 20 cycles until a steady voltammograms representative of a clean substrate is obtained (FIG. 6E). Then, electrodeposition of MoS.sub.2 was achieved across the gold electrochemical double layer potential range (0 to 1.4 V.sub.Ag/AgCl) at 100 mV/s for 20 cycles, in the three-cell configuration. Three Au microelectrodes were selectively and simultaneously modified in the process, i.e., by the oxidation of MoS.sub.2 through gold-thiol interaction using CV technique in the MoS.sub.2 solution (FIG. 6F).

WS.SUB.2 .Coated Microelectrodes (n.SUB.WS2.=3)

[0135] A protocol akin to the one described for the MoS.sub.2 electrodeposition was used to form WS.sub.2 coatings on three gold microelectrodes, which were simultaneously modified with the aid of the WS.sub.2 electrodeposition solution of Preparation 5. The microelectrodes were first cleaned in 0.1 mM sulfuric acid solution in the voltage range from −0.4 to 1.4V for 20 cycles. This allows the oxidation of different facets of polycrystalline gold. WS.sub.2 films were then produced using cyclic voltammetry (20 cycles, 0.1V/s rate, and voltage range: 0 to 1.4V) by the oxidation of WS.sub.2 and allowing the formation of gold-thiol interaction. FIGS. 6G and 6H show the CVs of the Au microelectrode cleaning (6G) and electrodeposition of WS.sub.2 on to the surface (6H) of three Au microelectrodes simultaneously.

Chitosan Coated Microelectrodes (n.SUB.chitosan.=3)

[0136] The chronopotentiometry technique was used to modify three gold microelectrodes in the arrayed chip with time t.sub.s=3 min and cathodic current j.sub.s=−4 μA/cm.sup.2. To this end, a two-electrode configuration was used, i.e., the counter electrode was shorted to reference terminal (see Preparation 6 for the electrodeposition solution). The electrodeposited microelectrodes were kept in buffer solution for 10 minutes to allow the weakly connected chitosan to be removed from the gold microelectrode surface. FIG. 6I shows the chronopotentiogram for the electrodeposition of chitosan film on the Au microelectrodes.

Chitosan-Carbon Nanotubes Coated Microelectrodes (n.SUB.chitosan-CNT.=3)

[0137] A protocol akin to the one described for the chitosan electrodeposition was used to form chitosan-CNT coatings on three gold microelectrodes (see Preparation 7 for the electrodeposition solution). FIG. 6J shows the chronopotentiogram for the electrodeposition of chitosan-CNT film on the Au microelectrodes.

[0138] FIG. 6K shows the optical image of bare and chitosan, chitosan-CNT, MoS.sub.2, WS.sub.2, r-GO, and Pt-black -modified gold microelectrodes along with on chip Ag/AgCl reference microelectrode in the multi-electrode array.

Example 2

Characterization of the Surface-Modified Gold Microelectrodes

SEM Analysis

[0139] An optical microscope (MX-50A, Olympus) was used to image the microelectrode before and after surface modifications. Electron micrograph images were obtained with a scanning electron microscope (JSM-7400F, JEOL Ltd.).

[0140] The presence of the materials in the multi-electrode array was examined using SEM (FIG. 7). FIGS. 7A and 7B show the bare gold microelectrode (7A; low resolution and 7B; high-resolution image). The high-resolution micrograph of the bare microelectrode shows the polycrystalline gold surface with its grain boundaries. FIGS. 7C and 7D show the electroplated gold microelectrode with Ag/AgCl (7C; low resolution and 7D; high-resolution image). The images reveal a porous structure of the metallic silver and silver chloride composite on to the gold microelectrode. The clusters of the Ag/AgCl can be clearly visualized in the high resolution micrograph. The presence of Pt-black was visualized in FIGS. 7E and 7F (7E: low resolution and 7F: high-resolution image). The electron micrographs indicate the porous assemblies of interconnected Pt-black crystals onto the gold microelectrode, which ensures an increased surface area. FIGS. 7G and 7H show the presence of r-GO flakes on the bare gold microelectrode (7G; low resolution and 7H; high-resolution image). The micrographs suggest the aggregation of the r-GO flakes onto the gold microelectrode. MoS.sub.2 nano flakes were visualized in FIGS. 7I and 7J (7I; low resolution and 7J; high resolution image). The electron micrographs reveal the presence of the gold microelectrode surface with MoS.sub.2 nano flakes. WS.sub.2 nano flakes were observed in FIGS. 7K and 7L (7K; low resolution and 7L; high resolution image). The micrograph shows the presence of WS.sub.2 nano flakes onto the gold microelectrode surface which was modified with WS.sub.2 monolayer solution. The electron micrograph in FIGS. 7M and 7N (7M; low resolution and 7N; high resolution image) show the surface morphology of the chitosan biopolymer on the bare gold microelectrodes whereas FIGS. 7) and 7P (7O; low resolution and 7P; high resolution image) reveal the presence of CNTs dispersed in biopolymer chitosan on the bare gold microelectrode.

Electrochemical Characterization

[0141] The detailed electrochemical characterization of the multi-electrode array was performed by recording the CVs in 10 mM PBS solution and in 5 mM ferricyanide/ferrocyanide solution. FIGS. 8B and 8C show the CVs recorded in 10 mM PBS solution using only Pt-black -modified microelectrode and the other material modifications, respectively, which is indicative of the background capacitive current that depends on the electroactive surface area of the bare and modified microelectrodes in the multi-electrode array. From FIGS. 8B and 8C, it is clear that the Pt-black -modified microelectrode has shown the highest background capacitive current as compared to all other modifications, which is due to the porous structure of the Pt-black that is clustered on to the gold microelectrode surface resulting in an increase in overall electroactive surface area of the microelectrode. FIG. 8D shows CVs recorded in 5 mM ferricyanide/ferrocyanide redox probe using the multi-electrode array, which shows the Nernstian behavior with I.sub.pa/I.sub.pc˜1 for all the microelectrodes (modified and bare) in the multi-electrode array (I.sub.pa/I.sub.pc=0.88 for Pt-black, I.sub.pa/I.sub.pc=0.92 for chitosan-CNT, I.sub.pa/I.sub.pc=0.95 for r-GO, I.sub.pa/I.sub.pc=0.96 for chitosan, I.sub.pa/I.sub.pc=0.96 for MoS.sub.2, I.sub.pa/I.sub.pc=0.98 for WS.sub.2, and I.sub.pa/I.sub.pc=0.98 for bare electrode). The Pt-black modified microelectrode has shown higher oxidation current as compared to microelectrode modifications (1.4, 1.8, 2.5, 6.9, 7.2, and 6.1 times higher than chitosan-CNT, r-GO, chitosan, MoS.sub.2, WS.sub.2 and bare) due to its high effective surface area and electrocatalytic activity.

Repeatability

[0142] For repeatability analysis, two microchips were coated with all the material modifications. Cyclic voltamograms in 10 mM PBS buffer before and after coating for 40 cycles (or more, depends on background signal stability) were recorded simultaneously. Following the PBS measurements, 20 cycles of voltamograms were recorded simultaneously in 5 mM ferrocyanide/ferricyanide before and after coating. After electrochemical measurements, thickness of the coatings were measured using AFM and profilometry (the thickness of the chitosan, chitosan-CNT film, Pt-black, and Ag/AgCl was measured by using a contact profilometry (Dektak-8, Veeco Ltd.). The thickness and the roughness of the r-GO film, MoS.sub.2 and WS.sub.2 were characterized using atomic force microscopy (MFP-3D-Bio, Asylum Research/Oxford Instruments) as they were very thin films as compared to other modifications. The same experiment was repeated five times; electrochemical measurement in PBS and ferrocyanide/ferricyanide followed by thickness measurements. The average peak oxidation current of 20 cycles and thickness for each modification was determined for each repetition and it was found that the fifth repetition has RSD value in thickness measurement of 2.7% for bare, 22% for chitosan, 17.98% for Chitosan-CNT, 4.2% for r-GO, 15.3% for Pt-black, 4.7% for MoS.sub.2, 4.3% for WS.sub.2, and 13.1% for Ag/AgCl. For 4.sup.th repeat the RSD in thickness measurement was below 10% for all the modification, indicating that the coating thickness was repeatable until four repeats.

Reproducibility

[0143] For the reproducibility analysis, five microchips were modified with all the modifications. For each chip the electrochemical measurement was performed in 10 Mm PBS followed by 5 mM ferricyanide/ferrocyanide solution. The average oxidation current for each coating was calculated and plotted against each coating. The RSD value of average oxidation current calculated for each modification for two chips (inter-chip; 6 microelectrode for each coating) and it was found below 10% (bare:6.1%, chitosan:8.6%, chitosan-CNT: 3.7, r-GO: 2.5%, Pt-black: 4.9%, MoS.sub.2: 4.8%, and WS.sub.2: 7.9%). The statistical t-test was performed which shows the p<0.05 for unpaired t-test. After electrochemical characterization of each chip, the thickness measurement was done using AFM and profilometry. The inter-chip RSD value in thickness measurement was also found below 10% (bare: 2.3%, chitosan: 7.6%, chitosan-CNT: 9.7, r-GO: 3.8%, Pt-black: 9.7%, MoS.sub.2: 7.0%, WS.sub.2: 7.1%, and Ag/AgCl: 6.7%). This shows that the multi-electrode array is reproducible in terms of electrochemical response as well as in thickness of each coating (FIGS. 8E and 8F).

Storage Stability

[0144] Five chips were modified with different materials and electrochemical measurement in 10 mM PBS, and in 5 mM ferro/ferricyanide solution were done simultaneously at day zero. The thickness measurement was also done at the same day. The chips were stored in separate bottles filled with PBS and kept in a dark and cool place. After one week, microchip #1 was taken out for the electrochemical and thickness measurement. Similarly, microchip #2 was characterized after two weeks, microchip #3 after three weeks, microchip #4 after four weeks, and microchip #5 after five weeks. The change in the electrochemical response and thickness was calculated as compared to day zero.

[0145] The thickness as found at the end of the five weeks test period was 99.08% for bare, 71.7% for chitosan, 77.8% for chitosan-CNT, 92.9% for r-GO, 78.8% for Pt-black, 85.7% for MOS.sub.2, 88.8% for WS.sub.2, and 60.86% for Ag/AgCl.

[0146] Raman spectrograms were measured by using a Raman spectrometer (LabRam HR, Horiba Ltd.).

Example 3

Detecting Clozapine in Blood Samples of Schizophrenia Patients Using the Multielectrode Array

Patients' Blood Processing SOP

[0147] Ten participants who had consented as part of a larger trial to have blood samples taken by finger-prick were recruited for the study. Participants in the study were diagnosed with schizophrenia or schizoaffective disorder and were eligible for clozapine treatment. Additionally, exclusion criteria included significant medical disorders or contraindications to the medication. Participants were generally titrated over the first month of treatment. Blood samples were collected prior to the start of clozapine treatment and at approximately 8-10 weeks after the start of clozapine treatment. Venous and capillary blood samples were collected at each visit. Approximately 8.5 mL of venous blood was collected in a red top BD (Becton, Dickinson and Company, Ltd.) collection tube using standard blood-drawing methods. Venous blood samples were centrifuged for 15 min at 1500 RCF and the serum was pipetted into a 3.5 mL transport tube and sent to Labcorp for clozapine blood serum analysis using LC-MS/MS technology.

[0148] Approximately 4 600 uL lavender top BD microtainers of capillary blood were collected via finger prick using 2 mm lancets and the Innovac Quick Draw capillary whole blood collection system. Whole blood from 2 of the microtainers were pipetted into a 1.5 mL freezer tube. Whole blood from the remaining 2 microtainers of capillary whole blood were centrifuged for 15 min at 1500 RCF and the serum was pipetted into 2 1.5 mL freezer tubes. Both the whole blood and plasma samples were stored in a −80° C. freezer prior to being sent to the laboratory for electrochemical analysis. The samples were thawed in an ice bucket before use.

Electrochemical Detection of Clozapine

[0149] The electrochemical signature of clozapine in whole blood, capillary plasma and capillary whole blood samples of schizophrenia patients was determined. 20 μL sample volume was used for all the sample measurements. The electrochemical data for each sample was recorded in two steps due to the limitation of the machine to record highly variable electrochemical signals. Three electrochemical signals for three Pt-black -modified microelectrode in the multi-electrode array was recorded simultaneously and then 18 electrochemical signals from rest of the modifications were recorded simultaneously. DPV technique was used (pulse width: 1 msec, pulse height: 55 mV, scan rate: 10 mV/sec, step height: 2 mV, Equilibration time: 10 sec, current range: 10 μA), to obtain relevant information on the redox reactions of clozapine in different solutions based on their standard potential. On chip Ag/AgCl as a reference microelectrode, and gold microelectrode in the center as a counter electrode, were used.

[0150] FIG. 9A and 9B show the smoothed DPVs recoded from 20 μL volume of capillary whole blood sample using the Pt-black -modified microelectrode only, and chitosan, chitosan-CNT, r-GO, MoS.sub.2, WS.sub.2-modified and bare microelectrodes. The CLZ concentrations that were obtained from the conventional methods were used to train and test the model performance. 80% of the electrochemical data set with full potential range was used to train the model and 20% of the data was used to test the model performance. The training model shows a PRESS value of 1.6 ng/mL (0.004 μM) for 3 number of factors to represent the variance of the electrochemical data set (FIG. 9C).

[0151] The linear regression analysis of training set for expected clozapine vs actual clozapine has shown a PCC value of 0.85 and LoD of 12.6±4.3 ng/mL (0.04±0.01) (FIG. 9D).

[0152] The trained model was further used to predict clozapine blood levels in patient samples and the predicted values were compared with the values obtained from commercial labs. The predicted values are 438±32.4 ng/mL (1.34±0.10) vs 450 ng/mL (1.4 μM) and 146±18.3 ng/mL (0.50±0.05) vs 131 ng/mL (0.40 μM). This shows a good degree of prediction of clozapine in patient's samples.

[0153] To obtain the results reported above, we have used the root mean square error (RMSE; Eq. 1), Pearson correlation coefficient (PCC; Eq.2) between the known and the estimated concentrations, in order to assess the linearity of the proposed model. We have used limit-of-detection (LoD; Eq. 3) in order to evaluate the sensing performance of the model.

[00004]$\begin{array}{cc}{\mathrm{RMSE}}_{\mathrm{test}}=\sqrt{\frac{1}{N_{\mathrm{test}}}{\left(C_{\mathrm{expected}}-C_{\mathrm{calculated}}\right)}^2}& \left(1\right)\end{array}$$\begin{array}{cc}\mathrm{PCC}=\frac{E\left[C_{\mathrm{expected}}-{\mu }_{\mathrm{expected}}\right]E\left[C_{\mathrm{estimated}}-{\mu }_{\mathrm{estimated}}\right]}{{\sigma }_{\mathrm{expected}}^2{\sigma }_{\mathrm{estimated}}^2}& \left(2\right)\end{array}$$\begin{array}{cc}\mathrm{LoD}=3*\mathrm{Sy}/x.fwdarw.\mathrm{Sy}/x=\sqrt{\frac{{.Math.}_i{\left(C_{\mathrm{expected}}-C_{\mathrm{estimated}}\right)}^2}{N_{\mathrm{test}}-2}}& \left(3\right)\end{array}$

[0154] Where C.sub.expected is known as the concentration in the solution and C.sub.estimated is the concentrations that was estimated by the regression model. E[x] describes the execution of the expected operation of specific vector, σ.sup.2.sub.expected and σ.sup.2.sub.estimated is the variance for the known concentration vector and the estimated ones, respectively. N.sub.test is the number of samples we used for model testing.