WO2001032303A1 - Batch fabrication of electrodes - Google Patents

Batch fabrication of electrodes Download PDF

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Publication number
WO2001032303A1
WO2001032303A1 PCT/US2000/030228 US0030228W WO0132303A1 WO 2001032303 A1 WO2001032303 A1 WO 2001032303A1 US 0030228 W US0030228 W US 0030228W WO 0132303 A1 WO0132303 A1 WO 0132303A1
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WO
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Prior art keywords
electrode
chamber
electrodes
surface
chlorine gas
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PCT/US2000/030228
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French (fr)
Inventor
Timothy D. Strong
Hal C. Cantor
Richard B. Brown
Robert W. Hower
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Advanced Sensor Technologies, Inc.
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    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by the preceding groups
    • G01N33/48Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/483Physical analysis of biological material
    • G01N33/4833Physical analysis of biological material of solid biological material, e.g. tissue samples, cell cultures
    • G01N33/4836Physical analysis of biological material of solid biological material, e.g. tissue samples, cell cultures using multielectrode arrays

Abstract

According to the present invention, there is provided a process for fabricating micro electrodes (10) by placing electrodes (10) in a vacuum chamber, purging the chamber of contaminated gases, flowing chlorine gas into the chamber and exciting ionized plasma into the chlorine gas, causing Cl- ions to form, accelerating these ions toward the surface where they react with the metal, thereby forming a metal salt on the surface of the electrodes (10). Also provided is an electrode made by placing the electrode in a vacuum chamber, purging the chamber of contaminants, gases, flow in chlorine gas into the chamber and exciting ionized plasma in the chlorine gas, causing Cl- ions to form, accelerating these ions toward the surface where they react with the metal, thereby forming a metal salt on the surface of the electrodes (10).

Description

BATCH FABRICATION OF ELECTRODES

BACKGROUND OF THE INVENTION

1. FIELD OF THE INVENTION

The present invention relates to methods of making electrodes for a sensor array. More specifically, the present invention relates to batch fabrication of electrodes in a low pressure chlorine plasma.

2. DESCRIPTION OF RELATED ART

Many standard chemical sensing devices rely on Ag/AgCI electrode as a reference contact to the solution under study and/or as the transducer of membrane potentials in ion selective electrodes. A properly formed Ag/AgCI electrode provides a nearly non-polarizable contact to the reference solution

(Bousse et al., 1986).

Conventional techniques used to fabricate Ag/AgCI electrodes, described in the literature, generally employ passing a small electrolyzing current through chlorine salt or dilute hydrochloric acid. The current is applied to an electrode coated with elemental Ag. AgCI is created in a layer on the electrode surface via a redox reaction with the solution containing Cl" ions (Bousse et al., 1986; Suzuki et al., 1998; Dendo et al., 1994).

The electrolysis technique suffers from several limitations. It is difficult to perform in a controlled manner on more than one electrode at a time. This can be quite time consuming in situations where arrays of Ag/AgCI electrodes are to be created. More importantly, it is extremely difficult to produce consistent results from electrode to electrode, requiring that each sensing site be calibrated independently to provide an accurate reference potential during use. The inconsistencies occur because the current flow, and therefore chloridization rate, is dependant on resistance of the electrical conducting line, surface area, and potential. All of these parameters are slightly different for every electrode, especially if several sites are chloridized at the same time. In addition, if the device uses on-chip readout electronics attached to the Ag/AgCI electrode, these electronics must take into account the electrolysis procedure to prevent damage, or must be designed to participate in the procedure itself, requiring special connections that must later be severed.

An additional technique for the chloridization of the electrodes is to use chemical chloridization. Chloridization can be accomplished in a FeCI3 solution. While this technique can be used to chloridize many sites at the same time, it is not selective, it produces an inferior Ag/AgCI layer, and it has a very fast chloridization rate that is difficult to control with the extremely thin Ag layer associated with planar electrodes. Additionally, small amounts of iron can be left as a contaminant on the sensor, affecting sensor transduction, as well as acting as a cellular toxin.

Analytical sensors are useful in chemistry and medicine to determine the presence and concentration of a biological and/or chemical analyte. Sensors are needed, for example, to monitor glucose in diabetic patients and lactate during critical care events. They are also needed to determine activity in cells, etc.

SUMMARY OF THE INVENTION

According to the present invention, there is provided a process for fabricating micro electrodes 10 by placing electrodes 10 in a vacuum chamber, purging the chamber of contaminant gases, flowing chlorine gas into the chamber and exciting ionized plasma into the chlorine gas, causing Cl" ions to form, accelerating these ions toward the surface where they react with the Ag metal, thereby forming a metal salt on the surface of the electrodes 10. Also provided is an electrode made by placing the electrode in a vacuum chamber, purging the chamber of contaminant gases, flowing chlorine gas into the chamber and exciting ionized plasma in the chlorine gas, causing Cl" ions to form, accelerating these ions toward the surface where they react with the Ag metal, thereby forming a metal salt on the surface of the electrodes 10.

DESCRIPTION OF THE DRAWINGS

Other advantages of the present invention are readily appreciated as the same becomes better understood by reference to the following detailed description when considered in connection with the accompanying drawing

wherein:

Figure 1 is an electron micrograph of the silver chloride surface showing fine grain size, large surface area, and totality of coverage;

Figure 2 is an x-ray spectrograph of the AgCI layer;

Figure 3 is an photo micrograph of the electrode of the present invention;

Figure 4 is a photograph of the wafer containing hundreds of sensors with thousands of electrodes; and

Figure 5 is a enlarged view of the center of the wafer depicted in Figure 4.

DETAILED DESCRIPTION OF THE INVENTION

Generally, the present invention provides a method of fabricating electrodes 10 and the electrodes 10 made by this process.

To remedy the problems with other chloridization techniques, a new technique for batch fabrication of electrodes 10 using a low pressure plasma which allows controllable wafer-level chloridation of all reference electrodes 10 at once, prior to separating the silicon wafer into individual chips.

In the preferred embodiment the electrodes 10 are Ag/AgCI electrodes 10. The plasma of this process is preferably a chlorine plasma, however other plasmas can be used as are known to those of skill in the art. The process uses a radio frequency (RF) plasma, produced at low pressure, to

add the necessary energy to decompose the chemical of the plasma, preferably chlorine, into reactive ions. The process includes the steps of placing macro electrodes, such as metal wire electrodes, and/or patterned wafers with micro electrodes (Figures 4 and 5) into a vacuum chamber. Then the chamber is carefully purged of contaminant gases and evacuated to high vacuum. Chlorine gas is flowed into the chamber while maintaining low pressure. An RF power source is used to excite an ionized plasma in the chlorine gas causing Cl" ions to form. The energized Cl" ions are accelerated toward the surface where they react with the exposed silver electrodes 10 forming AgCI on the surface (Figure 1).

The process replaces the conventionally employed electrolytic, non- electrolytic (i.e. FeCI3 method) and other chemical techniques for chloridation of silver-coated electrodes 10. The silver can be deposited onto wafers composed of silicon or other materials using any valid process, such as sputtering in an argon gas ambient. For testing purposes, 8000A of Ag was

applied to 200A of an adhesion promoter, titanium (Ti). The metal layers are applied to glass, ceramic, p- or n-type silicon wafers with 1.2μm of thermally

grown oxide, or silicon carbide. In production, the silver can be patterned into any shape desired via standard lift-off or etch procedures. The chloridation procedure is performed after patterning the silver.

Patterned wafers are placed in a standard RIE machine (for example the RIE machine from Plasma Therm Corporation). The chamber is evacuated to 2 x 10"5Torr. 40sccm N2 is flowed at 300mTorr for five minutes. The chamber is again evacuated to 2x10"5Torr. This process is intended to purge the chamber of any contaminant gasses that can affect the process,

and can be repeated as necessary to insure purity.

Next a mixture of Cl2 (or BCI3) and Argon (Ar) is flowed into the chamber. Test wafers are exposed to 40sccm of Cl2 or a mixture of 40 seem of Cl2 and 10sccm of Ar while pressure in the chamber is maintained at 300mTorr. An 18W or 25W plasma is then ignited in the chamber using the RIEs RF source. After ten minutes the chamber is purged, vented, and the silicon wafer removed.

Several parameters are precisely controllable using this technique, allowing fine manipulation of the final AgCI layer. The process pressure in the chamber can be altered: Initially, 300mTorr was employed, however, pressures from 100 mTorr to 700 mTorr have been utilized to control the reaction rate. The gas mixture can be precisely controlled using a mass flow control system, and/or other gases (N2 for example) can be utilized as diluents. Additionally, different power settings are available for the RF plasma. Higher power settings increase the reaction rate. Differing RF powers also have an effect on the crystal and physical structure of the AgCI layer produced. Finally different plasma exposure times provide a means of controlling the final thickness of the AgCI layer.

X-ray spectrographs were performed upon the AgCI layer to confirm composition (Figure 2). The X-ray spectrum of the RIE chlorinated silver is presented in Tables 1 and 2.

This technique provides a method to create Ag/AgCI reference electrodes 10 in a batch-wise process on whole wafers, chloridizing every reference electrode (of potentially thousands of sensors) at once, prior to separating the silicon wafer into individual chips. This new process generates AgCI layers that are more uniform, more controllable, and more repeatable than those produced utilizing electrolytic or chemical techniques. In addition the process can be carried out in standard reactive ion etch machines available in most clean room facilities.

In a preferred embodiment of the invention, cells are cultured upon a surface 6 according to known techniques. A layer of cells adhere to the respective surfaces of the microelectrode array 8 provided on the integrated device. As each of the microelectrodes 10 is driven with the applied voltage signal, a current flows between the microelectrodes 10 and the reference electrode. The impedance is determined from this current signal. Based on the impedance, various electrical characteristics of the cells which adhere to the surfaces of the microelectrodes 10 can be monitored. Such characteristics include the impedance of the individual cell (i.e., the combined cell membrane capacitance and conductance), the action potential parameters of the individual cell, the cell membrane capacitance, the cell membrane conductance, and the cell/substrate seal resistance.

The action potential parameters include, among others, action potential rate, action potential amplitude, and action potential shape and action potential power spectrum. The electrodes can also monitor the membrane

potential produced by the cell, such as neuronal action potentials.

It is a feature of the invention that each of the microelectrodes 10 is sufficiently small to enable monitoring of an individual cell and its cellular membrane.

Conventionally, the relative size of the reference electrode is large in comparison to the measuring electrodes 10 so that the measured impedance across each electrode and the reference electrode is dominated by the interface between the microelectrode and the cell, and the cell membrane impedance. In addition, the present invention utilizes microelectrodes 10 with a small surface such that the diameter of a cell is larger than the diameter of a microelectrode so that electrical characteristics related to individual cells and cell membranes can be resolved. The specific size of the microelectrodes 10 varies according to the specific application and size of the cells to be monitored. Preferably, the diameter of the microelectrodes 10 is less than or equal to one-half the diameter of the cell to be monitored, thereby permitting a given microelectrode to monitor an individual cell and cell membrane. For example, in one preferred embodiment, microelectrodes 10 with diameters of about 4 to 20 μm have been utilized. The combination of the relatively small size of the

microelectrodes 10, the relatively low impedance of the microelectrodes 10 (i.e., relative to cell membrane impedance) and the low noise characteristics of the signal detection components enables the present invention to monitor changes in the electrical characteristics of the cell, including changes in the capacitance and conductance of the cellular membrane. It also permits a variety of other techniques, described in greater detail below. These techniques include monitoring the action potential parameters of the cells; monitoring activation of voltage-gated ion channels; pharmaceutical screening and toxin detection; identifying particular cell/microelectrode junctions characterized by a higher degree of cell adherence to the microelectrode, and testing changes in cell adherence as adhesion promoting agents are employed.

The signal monitoring and processing means can include various devices for calculating the various cellular characteristics referred to above. For example, this element can include a personal computer configured to calculate the impedance between each microelectrode and the reference electrode based on the signals detected with the signal detecting means. By monitoring these values over time, the present invention can determine the various characteristics of individual cells which adhere to each microelectrode. The signal monitoring and processing means can be configured to perform a spectral analysis so as to monitor, in real time, characteristics such as changes in cell action potential parameters and cell impedance. It is appreciated that it is possible that more than one cell could be over a single microelectrode. The surface of a single microelectrode of the array 8 is exposed through a via in a passivation layer provided over a substrate. The microelectrode is driven with a signal source relative to the reference electrode.

The microelectrode has a known impedance Ze,ect. As known in the art, this value is described by a classical model of a metal in an electrolyte, and is calculated based on the material of the microelectrode or is measured separately prior to introduction of cells into the system. In series with the impedance of the microelectrode is the resistance of the solution Rsoln and the respective capacitance and conductive values associated with the microelectrode/cell junction. Parasitics to the substrate and parasitics to the passivation layer can be reduced by using a glass substrate, or can be factored out by normalizing.

Although often modeled as constants in prior art systems, the values of the capacitance and conductance associated with the cell actually vary in response to added toxins, pharmaceuticals and other substances, applied voltages, and changes in cellular morphology.

The impedances of the microelectrode, Zelect, and the reference electrode, Zref, can be determined in a variety of ways. These values, as well as the resistance of the solution, RS0|n, can be factored out, for example, by normalizing to data obtained prior to introduction of the cells to the system. Thus, by measuring the total impedance across each microelectrode and the reference electrode, it is possible to resolve impedances or changes in impedance relating to the cell motility, adhesion to the cell substrate (based on Rsea,), and changes in cellular membrane capacitance and conductance. It is further possible to monitor the activity of cellular ion channels and the action potential parameters of the cell as a result of their effects on the impedance measured between each microelectrode and the reference electrode or by actually monitoring the cell membrane potential. It is noted that the particular electrical characteristics of the cell, such as the capacitance and the conductance of the cellular membrane, are determined from the measured impedance by modeling the cell in accordance with known techniques. Various cell models can be used. For example, the cell can be modeled as a flat, circular "pancake" (i.e., as a disk). Other models can include a square or rectangular "pancake", a sphere, a cube, or a rectangular box.

In an embodiment of the invention, a homodyne detection technique is used to detect the signals resulting from the application of a signal voltage between each microelectrode and the reference electrode. A quadrature synthesizer is used to generate both sine and cosine signals of a programmed frequency and amplitude. The sine voltage signal is attenuated as needed and selectively applied to individual microelectrodes 10. The resulting signal is detected by a transimpedance stage which holds the large reference electrode at a reference potential (in this case, ground) via negative feedback. Automatic gain control is used to amplify the voltage output of the transimpedance stage. The amplified signal is multiplied in quadrature by the source signals and is low pass filtered to provide the real and imaginary components of the measurement. Quadrature multiplication allows for signal detection, for example, at the excitation frequency with high noise immunity. Known resistance values are used to calibrate the system for electrode impedance measurements.

In another embodiment, a quadrature synthesizer generates both a sine wave and cosine wave, A sin (ωt) and A cos (ωt). In this particular

example, the generated signals have a frequency of 1 kHz and an amplitude of 10 V peak to peak. Of course, it is understood that the present invention is not limited to these particular values. For example, a system has been constructed capable of applying signals from 100 Hz to 100 kHz. The sine wave is attenuated in this example, in a range from 0 to -80 dB. The attenuated signal is then selectively applied to particular microelectrodes 10. In this example, an analog multiplexer is used to apply the attenuated voltage signal to each of an array 8 of fifty-eight microelectrodes 10. Again, the invention is not limited to the number of microelectrodes 10 in the array 8 or to the components used for selective application of the signal to each of the microelectrodes 10.

The resulting current is detected by a transimpedance amplifier which maintains the large reference electrode at a virtual ground potential. The transimpedance amplifier outputs a signal, B sin (ω+φ). In this example, a

known resistance Rsense> 's applied across the input and output of the transimpedance amplifier.

The signal from the transimpedance amplifier is then amplified by an automatic gain control (AGC) amplifier having a variable gain Av. The signal is amplified in a range between 0 and 120 dB. The resulting signal,

Av B sin (ω+φ)

is multiplied in quadrature with mixers and then low pass filtered to obtain real and imaginary components X and Y. Thus, the signal from AGC amplifier is mixed with the signal A sin (ωt)

to obtain the following signal:

(Av AB/2)[cos φ-cos (2ωt+φ)]

The signal from the AGC amplifier is also mixed with the signal A cos (ωt) to

obtain the following signal: (Ay AB/2)[sin φ+sin (2ωt+φ)]

Thus, when low pass filtered, the following components remain:

X=(AV AB cos φ)/2 and

Y=(AV AB sin φ)/2 In one example, the system is calibrated with a known value resistance to obtain calibration values XCAL and YCAL. The magnitude |CAL| and phase φCAL of the calibration values are then determined as follows:

φCAL =arc tan [YCAL /XCAL ]|CAL|=[XCAL 2 +YCAL 2 f 2

Once the calibration values are obtained, measurements are taken with the

microelectrodes 10 and measurement values XMEAS and YMEAS are calculated as indicated above. Based on these values, the phase and magnitude for the respective measurements, φMEAS and |MEAS| are calculated as follow:

Φ EAS =arc tan[XMEAS /YMEAS ]|MEAS|=[XMEAS 2 +YMEAS 2 ]1/2 Given the respective values for XCAL, YCAL, XMEAS and YMEAS, one can divide the magnitude of the measured value |MEAS| by the magnitude of the calibration value |CAL| and then solve for the magnitude of the unknown impedance |ZUNKN0WN |. The phase of the unknown impedance ZUNKN0WN is

determined as follows: φzMEASCAL

The detected values for X and Y are sampled with an analog to digital (A/D) converter installed in a PC. In this example, a signal monitoring and processing means, such as a PC, receives the detected signal through an (A D) converter at a sample rate sufficient to read the detected sinusoidal signal. For example, the output from the AGC amplifier could be A/D converted and input to a PC. The signal monitoring and processing means obtains phase and magnitude values from the unknown impedance after directly converting the respective sinusoidal output for extraction of data. In this example, the signal monitoring and processing means provides the capability of performing a spectral analysis to monitor, in real time, characteristics such as changes in cell action potential parameters and cell impedance. By performing real time Fourier analysis, it is possible to monitor distortion caused by nonlinear electrode effects. Thus, it is a feature of the invention that the system can perform spectral analysis of the resulting signals in order to monitor changes in various characteristics in real time.

As described generally above, data obtained with the embodiment can be "normalized" to various conditions in order to monitor the characteristics of cells which adhere to the respective microelectrodes 10. For example, the membrane capacitance and conductance of individual cells which adhere to the microelectrodes 10 can be monitored, based on various models of the

cells Various other techniques are described herein

This embodiment provides high sensitivity and signal resolution for a large range of applications One characteristic of this embodiment is that the low pass filters have a characteristic settling time Where several electrodes 10 are monitored sequentially, this characteristic can introduce a minor delay It is possible to provide an even faster measurement cycle by heterodyning the detected signal to a higher frequency and then bandpass filtering to obtain phase and magnitude information Such a technique permits real time observation of extremely rapid variations in electrical characteristics of the cell, such as variations in transmembrane impedance (caused by opening and closing of ion channels)

A test signal oscillator generates sine and cosine signals, A sin (ωtest t)

and A cos (ωtest t) at a frequency ωtest The voltage sine signal is selectively

applied across each microelectrode and the reference electrode The resulting current is detected using transimpedance amplifier and AGC amplifier Mixers multiply the resulting signal B sin (ωtest t+φ) in quadrature by

sine and cosine signals generated by a local oscillator The signals produced by the local oscillator have an angular frequency ωL0, which is the sum of the

test frequency ωtest and an intermediate frequency ωIF As indicated above, the

respective outputs of mixers are represented, respectively, by (AB/2) cos [ωIF t+φ]-(AB/2) cos [(2ωtestIF)t+φ](AB/2) sin [ωIF t+φ]-(AB/2) sin [(2ωtestIF)t+φ]

These outputs are bandpass filtered at the intermediate frequency c%

and the filtered output detected using amplitude modulation detectors to provide real and imaginary components X=|(AB cos φ)/2| and Y=|(AB sin φ)/2|.

The algebraic sign of X and Y must be determined by a phase-sensitive detector, as is well known in the art. The unknown impedance between the microelectrode and the large reference electrode is then calculated in the manner described above.

It is appreciated that in addition to the exemplary embodiments discussed above, various other alternative embodiments are possible for impedance measurement. For example, measurement in the time domain can be used rather than homodyning to extract phase and magnitude information about each impedance. This can be accomplished, for example, by using a step function in place of the above-mentioned sinusoidal signal to drive each microelectrode. Further, the frequency of excitation and the sinusoidal input signal amplitude can be expanded beyond the exemplary ranges identified above.

Formed on the substrate is an insulation layer which electrically isolates a microelectrode array 8, such as that described above, from the substrate.

Between the substrate and the microelectrode is an optional thin film heater. The thin film heater is thermally coupled to, yet electrically insulated from the microelectrode array 8 which is disposed over the heater in this example. The thin film heater is used to maintain the microelectrode array 8 (and the electrolyte) at a substantially constant temperature (typically 37C). The thin film heater can be manufactured according to standard techniques known in the art, and provide more energy efficiency compared to non-

integrated heaters.

This example illustrates other optional features of the invention. Specifically, the integrated device includes various sensors which can be utilized alone or in combination with each other. For example, the sensors can include a pH sensor and/or a temperature sensor. As known in the art, the pH sensor can be constructed by deposition of a metal electrode (such as an iridium electrode) whose electrical potential varies with changes in pH or an Ag/AgCI electrode covered by a membrane with a hydronium ion selective ionophore, such as TDDA, Tridodecylamine. The inclusion of an integrated pH sensor not only provides for monitoring of pH changes caused by external factors, but also permits monitoring of pH changes caused by the cells which adhere to the integrated structure. Other ISE's or amperometric sensors can be employed to detect secretion of hormones, neurotransmitters, or other cellular metabolites.

Additionally, the integrated temperature sensor referred to above can advantageously provide closed loop temperature control, particularly when used in conjunction with the integrated heater. As with the other elements described above, the temperature sensor can be fabricated conveniently in accordance with standard microfabrication techniques. For example, layers of different metals can be deposited to form a thermocouple junction or the thermal coefficient of resistance (TCR) of the thin film heater could be used directly to monitor temperature. The output from this sensor could then be used via suitable external (or on-chip) circuits to selectively activate the thin film heater. Alternatively, it could be used to selectively activate non- integrated heating elements.

The sensors in the array 8 can be utilized in concert as: 1) ion selective electrodes 10 (ISEs) capable of monitoring a wide variety of important ions including electrolytes, stress hormones, CO2, local anesthetics, a variety of herbicides, heparin, medicinal drugs, lithium, etc., 2) amperometric electrodes 10 employing chronoamperometry and cyclic voltammetry for the detection of more complex molecules, such as hormones, neurotransmitters, neurotoxins and other environmental contaminants, and 3) electrodes 10 incorporating membranes with assay components that can be used to provide great sensitivity and selectivity, e.g. by immobilizing antibodies and/or enzymes on the surface of an ion-selective membrane and performing an enzyme-linked immunosorbent assay (ELISA) for example.

The microscopic sensor arrays are ideal for implementation of a micro- fluidic transdermal patch system in that they provide a large number of individual sensors, each of which can be encapsulated by a different membrane using an automated micro-screen printing device, such as the New Long LS-15TV, to confer sensitivity and specificity to individual biological ions and molecules of interest. Various modifications and alternative embodiments are apparent to those skilled in the art without departing from the invention. For example, the microelectrodes 10 and their respective interconnects and bond pads can comprise any biocompatible conductive substance, such as iridium, activated iridium, gold, platinum, polysilicon, aluminum, ITO, or TiW, bare or electroplated with platinum black.

Additionally, the substrate of the integrated device can be composed of a variety of materials, such as silicon, glass, metal, quartz, plastic, ceramic, polyethylene, or any other suitable type of polymer. It is noted that glass substrates have been found to provide reduced parasitic capacitance.

Other variations of the structure which are not essential to the underlying features of the invention include changes in the composition of the passivation layer. For example, devices made in accordance with the invention have utilized different passivation layers ranging from 0.5 to 5 μm in thickness. The passivation layer can comprise any suitable material, including low stress PECVD silicon nitride, silicon carbide, TEFLON™, polyimide, ceramic, photoresist, or any type of polymer or thermal plastic, or combination thereof.

Furthermore, the monolithic structures which include the microelectrode array 8 can be packaged in various configurations. According to one example, arrays 8 are fabricated on structures which are divided into individual 9χ9 mm chips. The chips are packaged in a standard 40-pin ceramic dual in-line package, or any other types of package which contains a suitable number of leads. In this example, the bondwires are encapsulated (for mechanical and electrical robustness) in a nontoxic low stress epoxy. Bondwire encapsulation can also be achieved with polyurethane, polyethylene, wax, TEFLON™, polyimide, etc. Of course this construction can be modified in many different ways. For example, instead of dicing the arrays 8 into multiple chips, the arrays 8 can remain on the same substrate to form multiwell plates, each containing an array 8 of electrodes 10. In such a configuration, Petri dishes are bonded to the substrate to define respective wells. The Petri dishes can be comprised of polystyrene, glass, polyethylene, TEFLON™, metal, or any other type of polymer. The Petri dishes can be bound to a chip formed in accordance with the invention using conventional materials and techniques, such as epoxy, polyurethane, wax, or thermoplastic, using thermal or ultrasonic processes. In this alternate construction, bondwire connections can be eliminated by use of a substrate configured to mate directly with a connector, such as an edge card connector or pads for a standard pin type connector. Of course, this requires the substrate area to be significantly larger than the active electrode 10 area.

The membrane capacitance, membrane conductance, cell/substrate separation and action potential parameters of a cell are significant markers regarding a cell's metabolic state, including general cellular health and ionic channel activity. The membrane potential, the voltage difference across a cell's lipid membrane, depends on the distribution of ionic charge. Generally, the distribution of ionic charge determines the electric potential, or voltage. For example, in a metallic conductor, the mobile particles carrying charge are electrons; in an aqueous solution, the mobile particles are ions such as Na+, K+, Cl", and Ca+2. In an aqueous solution, the number of positive and negative charges are normally balanced exactly, so that the net charge per unit volume is zero. An unbalanced excess of positive charges creates a region of high electrical potential, repelling other positive charges and attracting negative charges. An excess of negative charges repels other negative charges and attracts positive charge. When an accumulation of positive charges on one side of a membrane is balanced by an equal and opposite accumulation of negative charges on the other side of the membrane, a difference of electrical potential is set up between the two sides of the membrane.

Charge is carried back and forth across the cell membrane by small inorganic ions-chiefly Na+, K+, Cl", and Ca+2 , but these can traverse the lipid bilayer only by passing through special ion channels. When the ion channels open, the charge distribution shifts and the membrane potential changes. Of these ion channels, those whose permeability is regulated are the most significant; these are referred to as gated channels. Two classes of gated channels are of crucial importance: (1) voltage-gated channels, especially voltage-gated Na+ channels, which play the key role in the rapid changes in electrical energy by which an action potential is propagated along a nerve cell process; and (2) ligand-gated channels, which convert extracellular chemical signals into electrical signals, which play a central role in the operation of synapses. These two types of channels are not peculiar to neurons: they are also found in many other types of cells.

The present invention permits detection and monitoring of characteristics of a cell. As noted at the outset, these characteristics include cell impedance (cell membrane capacitance and conductance), action potential parameters, cell membrane capacitance, cell membrane conductance, cell/substrate seal resistance and/or the release of cellular products such as hormones, neurotransmitters or other cellular metabolites. These electrical and chemical characteristics in turn, correlate well with the metabolic state of the cell. The present invention, thus, provides an apparatus and method for monitoring cells and for monitoring the impact that an analyte has upon the metabolism of a cell. It is to be appreciated that such analytes include pharmaceutical agents, drugs, environmental factors, toxins, chemical agents, biological agents, viruses, and cellular adhesion promoters. Thus, the present invention is useful in screening and assaying broad classes of materials for their impact on cellular metabolism.

Other features of the invention become apparent in the course of the following descriptions of exemplary embodiments which are given for illustration of the invention and are not intended to be limiting thereof. Throughout this application, various publications, including United

States patents, are referenced by author and year and patents by number. Full citations for the publications are listed below. The disclosures of these publications and patents in their entireties are hereby incorporated by publications and patents in their entireties are hereby incorporated by reference into this application in order to more fully describe the state of the art to which this invention pertains

The invention has been descπbed in an illustrative manner, and it is to be understood that the terminology which has been used is intended to be in the nature of words of description rather than of limitation

Obviously, many modifications and variations of the present invention are possible in light of the above teachings It is, therefore, to be understood that within the scope of the appended claims, the invention can be practiced otherwise than as specifically described

Weight % by Element SiK C1K AgL

0.72 7.81 91.48

Table 1 : X-ray spectrograph weight percentage analysis of the AgCI layer.

Atomic % by Element SiK C1K AgL

2.34 20.13 77.53

Table 2: X-ray spectrograph atomic percentage analysis of the AgCI layer.

REFERENCES

1. L.J. Bousse, P. Bergveld, H.J.M. Geeraedts, Properties of Ag/AgCI Electrodes Fabricated with IC-compatible Technologies, Sensors and Actuators 9 (1986) 179-197.

2. H. Suzuki, A. Hiratsuka, S. Sasaki, I. Karube, Problems Associated with the Thin-film Ag/AgCI Reference Electrode and a Novel Structure with Improved Durability, Sensors and Actuators B 46 (1998) 104-113.

3. I. Dendo, N.F. Sheppard, M. Eden, G. Kantor, Precision Silver/Silver Chloride Electrodes, Proceedings of the 16th Annual International Conference of the IEEE Engineering in Medicine and Biology Society 2 (1994) 810-811.

Claims

CLAIMSWhat is claimed is:
1. A process for fabricating microelectrodes by: placing electrodes having a surface in a vacuum chamber; purging chamber of contaminant gases; flowing chlorine gas into the chamber; exciting an ionized plasma in the chlorine gas causing Cl" ions to form and forming a salt on the surface of the electrodes.
2. The process according to claim 1 , wherein said excitation step includes using a power source to excite the ionized plasma.
3. The process according to claim 1 , wherein said flowing step includes flowing said chlorine gas into the chamber while maintaining low pressure.
4. The process according to claim 1 , wherein said exciting step further includes the step of accelerating the Cl" ions toward the surface where the ions react with the silver metal.
5. An electrode made by: placing said electrode having a surface in a vacuum chamber; purging said chamber of contaminant gases; flowing chlorine gas into said chamber; exciting an ionized plasma in the chlorine gas causing Cl" ions to form and forming a metal salt on the surface of the electrodes.
6. The electrode according to claim 5, wherein said electrode is a silver electrode.
7. The electrode according to claim 6, wherein AgCI is formed on the surface of said electrode.
8. The electrode according to claim 5, said excitation step includes using a power source to excite said ionized plasma.
9. The electrode according to claim 8, wherein said power source is a RF power source.
10. The electrode according to claim 5, wherein said flowing step includes flowing said chlorine gas is flowed into said chamber while maintaining low pressure.
11. The electrode according to claim 5, wherein said electrode is constructed on a substrate.
12. The electrode according to claim 11 , wherein said substrate is a wafer.
13. The electrode according to claim 5, wherein said exciting step further includes the step of accelerating the Cl" ions toward the surface where the ions react with the silver metal.
14. The electrode according to claim 5, wherein said electrode is made on a material selected from the group consisting essentially of glass, silicon carbide, ceramic, silicon and plastic.
PCT/US2000/030228 1999-11-02 2000-11-02 Batch fabrication of electrodes WO2001032303A1 (en)

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Publication number Priority date Publication date Assignee Title
WO2002035221A1 (en) * 2000-10-27 2002-05-02 Uutech Limited Method for chlorine plasma modification of silver electrodes
WO2010023569A1 (en) * 2008-08-25 2010-03-04 Nxp B.V. Reducing capacitive charging in electronic devices
US9322103B2 (en) 2010-08-06 2016-04-26 Microchips Biotech, Inc. Biosensor membrane composition, biosensor, and methods for making same

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JPS59182357A (en) * 1983-03-31 1984-10-17 Shimadzu Corp Ion sensor
US5200053A (en) * 1987-11-24 1993-04-06 Terumo Kabushiki Kaisha Reference electrode

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US4062750A (en) * 1974-12-18 1977-12-13 James Francis Butler Thin film electrochemical electrode and cell
JPS59182357A (en) * 1983-03-31 1984-10-17 Shimadzu Corp Ion sensor
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Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2002035221A1 (en) * 2000-10-27 2002-05-02 Uutech Limited Method for chlorine plasma modification of silver electrodes
WO2010023569A1 (en) * 2008-08-25 2010-03-04 Nxp B.V. Reducing capacitive charging in electronic devices
US9006738B2 (en) 2008-08-25 2015-04-14 Nxp, B.V. Reducing capacitive charging in electronic devices
US9322103B2 (en) 2010-08-06 2016-04-26 Microchips Biotech, Inc. Biosensor membrane composition, biosensor, and methods for making same

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