United States Patent [19]
`Hill et al.
`
`US005820551A
`[11] Patent Number:
`[45] Date of Patent:
`
`5,820,551
`*Oct. 13, 1998
`
`[54] STRIP ELECTRODE WITH SCREEN
`PRINTING
`
`[56]
`
`References Cited
`
`U.S. PATENT DOCUMENTS
`
`[76] Inventors: Hugh Allen Oliver Hill, Nine Clover
`Close, Oxford OX2 9JH; Irving John
`Higgins, Cotswold, Graze Hill,
`Ravensden, Bedford MK44 2TM;
`James Michael McCann, 4 Ash Grove,
`Headington, Oxford, all of Great
`Britain; Graham Davis, 1504 Fox Run
`Dr., Plainsborough, N]. 08536
`
`[ * ]
`
`Notice:
`
`The term of this patent shall not extend
`beyond the expiration date of Pat. No.
`5,682,884.
`
`3,352,762 11/1967 Weiner.
`3,542,662 11/1970 Hicks 6161..
`3,838,033
`9/1974 Mindt et al. .
`
`(List continued on next page.)
`
`FOREIGN PATENT DOCUMENTS
`
`0305 03A1
`078636A1
`
`6/1981 European Pat. Off. .
`5/1983 European Pat. Off. .
`
`(List continued on next page.)
`
`OTHER PUBLICATIONS
`
`[21] Appl. No.: 470,352
`
`[22]
`
`Filed:
`
`Jun. 6, 1995
`
`Related U.S. Application Data
`
`[63] Continuation of Ser. No. 281,131, Jul. 27, 1994, Pat. No.
`5,682,884, which is a continuation of Ser. No. 888,264, May
`22, 1993, abandoned, which is a continuation of Ser. No.
`768,359, Sep. 30, 1991, abandoned, which is a continuation
`of Ser. No. 634,968, Jan. 7, 1991, abandoned, which is a
`continuation of Ser. No. 2,120, Jan. 12, 1987, abandoned,
`which is a continuation-in-part of Ser. No. 607,599, May 7,
`1984, abandoned.
`
`[30]
`
`Foreign Application Priority Data
`
`May 5, 1983 [GB]
`May 5, 1983 [GB]
`Jun. 6, 1983 [GB]
`Dec. 16, 1983 [GB]
`Jan. 11, 1984 [GB]
`Feb. 29, 1984 [GB]
`Aug. 29, 1984 [GB]
`
`United Kingdom ................. .. 8312261
`United Kingdom
`8312262
`United Kingdom
`8323799
`United Kingdom
`8333644
`United Kingdom
`8400650
`United Kingdom
`8405262
`United Kingdom ................. .. 8405263
`
`[51] Int. Cl.6 ...................................................... .. A6113 5/05
`
`[52] U.S. Cl. ........................ .. 600/347; 600/345; 600/365;
`204/403; 204/406; 204/407; 204/435
`[58] Field of Search ................................... .. 128/633, 634,
`128/637, 692, 639; 204/1531, 153.12,
`402, 403, 406, 407, 435; 600/309, 345,
`347, 348, 365, 368
`
`AizaWa et al., “Amperometric Determination of Human
`Chorionic Gonadotropin by Membrane—Bound Antibody,”
`Enzyme Immunosensor 22—28, 1978.
`
`(List continued on next page.)
`
`Primary Examiner—David M. Shay
`
`[57]
`
`ABSTRACT
`
`A sensor system that detects a current representative of a
`compound in a liquid mixture features a tWo-electrode strip
`adapted for releasable attachment to signal readout circuitry.
`The strip comprises an elongated support (preferably ?at)
`adapted for releasable attachment to the readout circuitry; a
`?rst conductor and a second conductor each extend along the
`support and comprise means for connection to the circuitry.
`An active electrode, positioned to contact the liquid mixture
`and the ?rst conductor, comprises a single layer deposit of
`an enzyme capable of catalyzing a reaction involving the
`compound, a conductive material and an electron mediator,
`capable of transferring electrons betWeen the enzyme
`catalyzed reaction and the ?rst conductor. A reference elec
`trode is positioned to contact the mixture and the second
`conductor. The system includes circuitry adapted to provide
`an electrical signal representative of the current. The tWo
`electrode strip is manufactured, e.g., by screen printing an
`admixture of an enzyme, a conductive material, and a
`mediator as a single layer onto the substrate.
`
`9 Claims, 6 Drawing Sheets
`
`LIFESCAN SCOTLAND LTD. EXHIBIT 2007
`
`

`
`5,820,551
`Page 2
`
`US. PATENT DOCUMENTS
`
`OTHER PUBLICATIONS
`
`Brown, Jr. .
`Kowarski .
`Reimes et al. .
`
`Toida ................................ .. 350/339 R
`AfromoWitZ et al.
`128/642
`Rolfe ..................................... .. 128/635
`Goller et al. .
`
`Johnson .
`Nakamura et al. .
`
`Pace .
`Frank et al. .
`Takinishi et al. .
`Nakashima et al. .
`
`Johnson .
`Nakamura et al. .
`Junter et al. .
`Brown et al. .
`Burns et al. .
`
`Pace .
`
`Slama .
`
`8/1975
`2/1977
`4/1977
`1/1978
`1/1979
`12/1979
`1/1980
`8/1980
`9/1980
`9/1980
`10/1980
`6/1982
`10/1982
`10/1982
`3/1983
`6/1983
`7/1983
`5/1984
`6/1984
`9/1984
`12/1984
`12/1984
`10/1985
`6/1986
`12/1987
`11/1997
`
`3,900,382
`4,008,717
`4,019,966
`4,068,923
`4,133,735
`4,176,659
`4,185,131
`4,216,245
`4,224,125
`4,225,410
`4,229,490
`4,334,542
`4,352,884
`4,356,074
`4,376,689
`4,390,620
`4,394,070
`4,449,529
`4,454,007
`4,469,110
`4,488,556
`4,490,464
`4,545,382
`4,592,824
`4,711,245
`5,682,884
`FOREIGN PATENT DOCUMENTS
`
`Ho ......................................... .. 128/635
`Gorton et al. .
`Higgins et al. ....................... .. 128/635
`Smith et al. .
`Higgins et al. .
`Hill et al. .............................. .. 600/368
`
`127958A2
`152541A1
`0351891A2
`2127142
`54-130196
`60-29658
`61-161443
`61-270652
`827023
`1318815
`2175400
`WO 84/03562
`
`12/1984
`8/1985
`1/1990
`6/1971
`3/1978
`7/1983
`7/1986
`11/1986
`5/1981
`5/1973
`5/1986
`9/1984
`
`European Pat. Off. .
`European Pat. Off. .
`European Pat. Off. .
`Germany .
`Japan .
`Japan .
`Japan .
`Japan .
`
`U.S.S.R. .
`United Kingdom .
`United Kingdom .
`
`WIPO .
`
`Bertrand et al., “Multipurpose Electrode With Different
`EnZyme Systems Bound to Collagen Films,” Analytics
`Chemics Acts 126:23—34, 1981.
`Brodnick et al., “Laminated Electrodes for Biopotential
`Studies”, IEEE Transactions on Biomedical Engineering
`BME—25:479—481, 1978.
`Hill, “The Exploitation of the Electrochemistry of Proteins”,
`Biochemical Society Transactions 11:453—455 1983.
`Koryta et al., “Electroanalytical Methods Used in Medicine
`and Biology”, Medical and Biological Applications of Elec
`trochemical Devices 7—11, 1980.
`Liu et al., “Fabrication of Miniature PO2 and pH Sensors
`Using Microelectronic Techniques”, Diabetes Care
`5:275—275—277, 1982.
`Liu et al., “Miniature Multiple Cathode Dissolved Oxygen
`Sensor for Marine Science Applications”, Marine Technol
`ogy 16:468—472, 1980.
`May et al., “A Tantalum—on—Sapphire Microelectrode
`Array”, IEEE Transactions on Electron Devices
`ED—26:1932—1939, 1979.
`Oehme, “Chemische Sensoren,” VieWeg Verlag, p. 76, 1991
`no translation.
`Pickard et al., “Flexible Printed—Circuit Probe for Electro
`physiology”, Med. & Biol. Eng. & Comput. 17:261—267,
`1979.
`Turner et al., “Applications of Electron Transfer BetWeen
`Biological Systems and Electrodes”, Biochemical Society
`Transactions 11:445—448, 1983.
`EnZyme Electrode”, Nature
`Updike et
`al., “The
`214:986—988, 1967.
`Technical Note—“A Microelectrode Ampli?er With Novel
`Frequency Compensation”, Med. & Biol. Engng.
`9:557—561, 1971.
`
`

`
`U.S. Patent
`
`0a. 13, 1998
`
`Sheet 1 of6
`
`5,820,551
`
`FIG. 1
`
`

`
`U.S. Patent
`
`Oct. 13, 1998
`
`Sheet 2 0f 6
`
`5,820,551
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`{/24
`
`F165
`
`F167
`as
`
`//
`
`40
`
`43
`
`39
`
`42
`
`A0
`A2
`
`

`
`U.S. Patent
`
`0a. 13, 1998
`
`Sheet 3 of6
`
`5,820,551
`
`MA
`
`>FIG. 8
`
`0
`+200
`E / mV vs.SCE.
`
`+ 400
`
`FIG. 9a
`
`FIG. 9b
`
`2
`
`L
`
`'7
`
`9
`
`6
`
`

`
`U.S. Patent
`
`0a. 13, 1998
`
`Sheet 4 0f 6
`
`5,820,551
`
`705
`
`'// 705
`
`REFERENCE
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`
`'04
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`VOLTAGE
`BUFFER
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`UIVIUER
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`
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`
`

`
`U.S. Patent
`
`Oct. 13, 1998
`
`Sheet 5 of6
`
`5,820,551
`
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`
`ma3w2
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`

`
`U.S. Patent
`
`0a. 13, 1998
`
`Sheet 6 of6
`
`5,820,551
`
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`
`1
`STRIP ELECTRODE WITH SCREEN
`PRINTING
`
`CROSS-REFERENCE TO RELATED
`APPLICATIONS
`
`This is a continuation of application Ser. No. 08/281,131
`?led on Jul. 27, 1994 Which is a continuation of Ser. No.
`07/888,264, ?led May 22, 1993; Which is a continuation of
`Ser. No. 07/768,359, ?led Sep. 30, 1991; Which is a con
`tinuation of Ser. No. 07/634,968, ?led Jan. 7, 1991; Which is
`a continuation of Ser. No. 07/002,120, ?led Jan. 12, 1987;
`Which is a continuation-in-part of Ser. No. 06/607,599, ?led
`May 7, 1984, noW abandoned. The following US. patent
`applications ?led May 7, 1984 and oWned by applicants’
`assignee, Medisense, Inc. are hereby incorporated by refer
`ence: Sensor For Components Of A Liquid Mixture (U.S.
`Ser. No. 607,699); Assay Techniques Utilizing Speci?c
`Binding Agents (U.S. Ser. No. 607,695); Assay Systems
`Using More Than One Enzyme (U.S. Ser. No. 607,698);
`Assay Systems Utilizing More Than One Enzyme; (U.S. Ser.
`No. 607,607).
`
`BACKGROUND OF THE INVENTION
`
`This invention relates to enzymatic sensor electrodes and
`their combination With reference electrodes to detect a
`compound in a liquid mixture.
`Various electrochemical sensors are knoWn Which employ
`enzymes to sense the presence of a compound that serves as
`an enzyme substrate. As just one example, Nakamura US.
`Pat. No. 4,224,125 discloses an enzyme electrode system in
`Which an enzyme, such as glucose oxidase, is used to sense
`glucose. A redox compound is used to accept electrons from
`the enzyme. For example, at 10:21—49, Nakamura discloses
`(FIG. 5) press molding to the electrode a mixture of glucose
`oxidase cross-linked by gluteraldehyde and a ?uorocarbon
`polymer poWder together With a cation exchange resin
`containing potassium ferricyanide. Nakamura’s electrode
`system (FIG. 2) consists of three electrodes: an enzyme
`electrode 7, a reference electrode 8, and a counter electrode
`10.
`In another example, Pace US. Pat. No. 4,225,410 dis
`closes a multi-layer enzyme sensor; for example sensor 14b
`(FIG. 7b) measures levels of lactate dehydrogenase. NAD+
`is generated at a fourth electrode 82, and the enzymatic
`reaction converts it to NADH Which is sensed at monitoring
`electrode 84 by undisclosed means. A barrier/counter elec
`trode 80 and a reference electrode 85 are used in conjunction
`With monitoring electrode 84.
`
`10
`
`15
`
`20
`
`25
`
`30
`
`35
`
`40
`
`45
`
`SUMMARY OF THE INVENTION
`
`55
`
`One aspect of the invention generally features a tWo
`electrode strip for releasable attachment to signal readout
`circuitry, forming a sensor system that detects a current
`representative of a compound in a liquid mixture. The strip
`comprises an elongated support (preferably ?at) adapted for
`releasable attachment to the readout circuitry; a ?rst con
`ductor and a second conductor each extend along the support
`and comprise means for connection to the circuitry. An
`active electrode, positioned to contact the liquid mixture and
`the ?rst conductor, comprises a deposit of an enzyme
`capable of catalyzing a reaction involving the compound.
`Electrons are transferred betWeen the enzyme-catalyzed
`reaction and the ?rst conductor to create the current. A
`65
`reference electrode is positioned to contact the mixture and
`the second conductor.
`
`60
`
`5,820,551
`
`2
`The preferred embodiment of the strip includes the fol
`loWing features. An electron mediator (most preferably a
`ferrocene) is included in the active electrode deposit to effect
`the electron transfer. The compound being detected is
`glucose, and the enzyme is glucose oxidase or glucose
`dehydrogenase. The active electrode and the reference elec
`trode are coatings applied to the elongated support, eg the
`active electrode is formed by printing (e.g. screen printing)
`an ink comprising a conductive compound, the enzyme and
`the mediator, and the reference electrode is also formed by
`screen printing. The means for connecting to the readout
`circuit are positioned toWard one end of the elongated
`support, and the electrodes are positioned remote from that
`end.
`In a second aspect, the invention features apparatus that
`includes the above-described strip and circuitry adapted to
`provide an electrical signal representative of the current. The
`read-out circuitry comprises an ampli?er to amplify the
`current, a loW-pass ?lter connected to the ampli?er output,
`a current-to-voltage converter connected to the ?lter output,
`and a means to display a value representative of the output
`of the converter. The apparatus also includes means to
`maintain a substantially constant bias voltage across the
`electrodes during current measurement, even Without a
`separate voltage reference in contact With the mixture.
`In a third aspect, the invention features screen printing the
`enzyme onto a substrate to form an enzymatic sensing
`electrode. The ink used for screen printing includes a liquid
`vehicle, a suspension of conductive material, and the
`enzyme. Preferably, it also includes a mediator capable of
`transferring electrons betWeen the enzymatic reaction and a
`conductor on the substrate. Also preferably, the substrate is
`a ?exible, high-dielectric polymeric substance, such as poly
`vinyl choride, polyester, or polycarbonate.
`The invention enables a very small, inexpensively
`manufactured, disposable electrode strip that provides an
`accurate electronic readout of the target compound. In
`particular, the active electrode is sized to be covered by the
`smear of blood produced from a drop of blood (even a
`non-expressed drop) generated from a needle-prick at a
`bodily extremity, and the reference electrode is sized and
`spaced from the active electrode a distance such that the
`reference electrode is covered by the same smear of blood.
`Other features and advantages of the invention Will be
`apparent from the folloWing description of the preferred
`embodiment.
`
`DESCRIPTION OF THE PREFERRED
`EMBODIMENT
`We ?rst brie?y describe the draWings.
`DraWings
`FIG. 1 is a front vieW of a strip-supported electrode
`con?guration;
`FIG. 2 is a back vieW of the combination shoWn in FIG.
`1;
`FIG. 3 shoWs an alternative strip-supported electrode;
`FIG. 4 shoWs a strip-supported electrode Which is a
`variant of FIG. 3;
`FIG. 5 shoWs a modi?ed connection of the strip electrode
`of FIGS. 3 and 4;
`FIG. 6 shoWs a further alternative supported electrode;
`FIG. 7 shoWs a combination of tWo electrode supports;
`FIG. 8 shoWs diagrammatically the electrical performance
`of electrodes as described in the above Figures, With especial
`reference to FIGS. 6 and 7;
`
`

`
`5,820,551
`
`3
`FIG. 9a and 9b are general diagrammatic side views of a
`pen-like portable holder, of particular utility for the elec
`trodes shown in FIGS. 3, 4 and 5, having an assembly of
`circuitry and having a read-out WindoW;
`FIG. 10 shoWs a schematic diagram of one form of
`electrical circuitry for use With the electrodes and equipment
`of the present invention;
`FIG. 11 shoWs a more elaborated circuit diagram for use
`in the embodiment of FIG. 10;
`FIG. 12 shoWs a schematic diagram of an alternative
`embodiment of electrical circuitry; and
`FIG. 13 shoWs the more elaborated circuit diagram of a
`yet further embodiment of circuitry.
`
`-continued
`
`Substrate
`
`Enzyme
`
`PQQ Enzymes
`
`Glucose Dehydrogenase
`Methanol Dehydrogenase
`
`Methylamine Dehydrogenase
`Haem-Containing Enzymes
`
`Lactate Dehydrogenase
`(Yeast Cytochrome b2)
`Horse-Radish Peroxidase
`Yeast Cytochrome c
`Peroxidase
`Metallo?avoproteins
`
`Carbon Monoxide
`Oxidoreductase
`Cup ropro teins
`
`Glucose
`Methanol and Other
`Alkanols
`Methylamine
`
`Lactate
`
`Hydrogen Peroxide
`
`Hydrogen Peroxide
`
`Carbon Monoxide
`
`Galactose Oxidase
`
`Galactose
`
`The strip electrode has the folloWing design criteria. The
`electrodes on the strip should be as small as possible and the
`strip should preferably be disposable. The strip should be
`elongate for ready handling as an electrode for ready assem
`bly to equipment on the one hand and contact With the
`sample on the other. It must be sensitively manipulable. It
`must carry, prior to assembly or in the assembled structure,
`the reference electrode as Well as the ‘sensitive’ electrode, in
`spaced non-contiguous relationship.
`The invention is particularly useful for selective
`detection, measurement or monitoring of a given dissolved
`substrate in a mixture of dissolved substrates.
`The elongate support could be a rod or tube, but conve
`niently it comprises a ?at strip.
`The active electrode is preferably formed of carbon e.g.,
`a ?lter paper containing carbon. We have also found that
`carbon foil e.g., as available under the Trade Marks “GRA
`PHOIL” or “PAPYEX” is a valuable electrode material. The
`enzyme thereon can in theory be any enzyme, e.g., those
`listed in US. Ser. No. 607,699, and itemized above but the
`use of glucose oxidase or dehydrogenase, e.g., the bacterial
`glucose dehydrogenase from Acinetobacter calcoaceticus is
`particularly valuable. Any suitable mediator compound can
`be used, but ferrocene or ferrocene derivatives (expecially
`1,1‘-dimethylferrocene) are greatly to be preferred.
`By Way of example only, carbon foil can be glued to the
`strip; 1,1‘-dimethylferrocene mediator can be deposited on
`the surface of the foil by evaporation of a toluene solution;
`and enzyme can be bonded to the surface by the use of
`1-cyclohexyl-3-(2-morpholinoethyl) carbodiimide metho-p
`toluene sulphonate (referred to beloW as “carbodiimide”).
`The reference electrode can be any convenient reference
`electrode. We have found it useful to provide adjacent but
`not contiguous to the ?rst electrode, a ?at layer of silver and
`to convert the surface thereof to silver chloride so as to give
`an Ag/AgCl reference electrode.
`Typically, the electrical connections can be Wires Which
`extend doWn, and are preferably adhered to, the strip, and
`make electrical contact each With its respective electrode.
`The readout means is preferably a digital indicator suit
`ably connected to a dedicated potentiostat Which poises the
`carbon electrode potential at e.g. +150 mV vs. Ag/AgCl for
`a glucose system. The current ?oWing is then proportional to
`glucose concentration.
`In a particular valuable version of this type of sensor, it
`comprises (a) a ?at ?rst electrode area of knoWn area small
`
`10
`
`15
`
`20
`
`25
`
`30
`
`35
`
`40
`
`45
`
`55
`
`60
`
`65
`
`ELECTRODE STRUCTURE
`
`In general, the strip electrode of the invention comprises
`a conductive electrode coated With a mixture, or layers, of a
`catalytically active enzyme and a mediator compound and
`usually further coated With a retaining permeable mem
`brane. When such a coated electrode is contacted With a
`substrate containing a species for Which the enzyme exerts
`a catalytic effect, the mediator compound transfers charge to
`the electrode and this can be used to give a readout signal,
`against a standard electrode, correlated With the concentra
`tion of the said species, even in the presence of other species
`since enzymes are typically highly selective in their catalytic
`action. Commonly oWned Ser. No. 607,699 generally
`describes methods of coating a conductive electrode With
`enzyme and mediator; that application is hereby incorpo
`rated by reference. The mediator compounds described in
`Ser. No. 607,699 include polyviologens, ?uoranil and chlo
`ranil. HoWever, the preferred mediator compounds are met
`allocene compounds, and in particular the ferrocenes
`(biscyclopentadienyl iron and its derivatives).
`The particular advantages of ferrocenes are as folloWs: (a)
`a Wide range of redox potentials accessible through substi
`tution of the cylopentadienyl rings (b) functionalization of
`the rings, e.g., to confer solubility or chemical linkability to
`other such rings or other system components (c) electro
`chemically reversible one-electron redox properties (d)
`pH-independent redox potential and (e) sloW autooxidation
`of the reduced form.
`The ferrocene structure may be modi?ed by substitution
`on the rings, and/or by association or polymerization, Which
`modi?cations affect the physical, chemical and electrical
`behavior so that optimization of a particular sensor electrode
`material is possible. In general use, the compound 1,1‘
`dimethylferrocene is a valuable mediator. The particular
`enzyme employed may be selected from a range of enzymes
`including the folloWing:
`
`Enzyme
`
`Flavo-Proteins
`
`Pyruvate Oxidase
`Amino Acids Oxidase
`Aldehyde Oxidase
`Xanthine Oxidase
`Glucose Oxidase
`Glycollate Oxidase
`Sarcosine Oxidase
`Lactate Oxidase
`Glutathione Reductase
`Lipoamide Dehydrogenase
`
`Substrate
`
`Pyruvate
`
`Aldehydes
`Xanthines
`Glucose
`Glycollate
`Sarcosine
`Lactate
`NAD(P)H
`NADH
`
`

`
`5,820,551
`
`5
`enough to be completely coverable by the smear of blood
`produced from a non-expressed drop of blood generated
`from a needle-prick at a bodily extremity, (b) a reference
`electrode area on the same surface separate from but suffi
`ciently close to the sensitive electrode area that the said
`blood smear also reaches the reference electrode to establish
`electrical communication, and (c) conductive elements
`extending separately along the same surface of, and thus
`insulated from the elongate support member, communicat
`ing one With each electrode for connection to signal readout
`means attachable to one end of the member.
`The area of the ?rst (i.e. sensitive or active) electrode is
`generally substantially square; it may be rectangular or
`otherWise shaped, but in any case usually Will correspond in
`area to a square of 5 mm edge length, or beloW e.g., from 2
`to 4 mm.
`For convenience, this document Will refer hereinafter to
`blood-glucose-measuring equipment as being typical but not
`limitative of equipment With Which the present invention is
`concerned.
`Diabetic subjects need to measure their glucose levels
`frequently. Hitherto, a common method carried out by the
`subject personally is calorimetric test using a blood or urine
`sample Which is applied over a surface area containing a
`color-reactive detector chemical, adjacent to a comparison
`area, to give a color change Which is compared With a chart
`of color values as an approximate measure of glucose level.
`There are hoWever, defects in this method. Firstly, calo
`rimetric changes are quantitatively dif?cult to assess, espe
`cially if the patient has impaired vision as a result of the
`diabetic condition. Indeed, because of this problem expen
`sive automatic color comparison equipment may need to be
`purchased by some subjects for interpreting the test results.
`Secondly, the blood test, While inherently more accurate
`than a urine test, needs a large enough sample to cover the
`test surface. Thirdly, it requires the patient to time the color
`development accurately. Since blood samples, on a self
`treatment basis, are taken from body extremities (?ngers,
`toes, earlobes), they are normally not large enough When
`obtained by a simple needle-prick, and must in fact be
`expressed i.e., squeeZed or massaged out to form a larger
`drop. Progressively, the tissue of the extremities becomes
`scarred and coarsened by such treatment to an extent
`Whereby ?nding fresh testing sites presents a problem.
`In order to embody the invention on a home-diagnostic
`basis, a main object of the present invention in one aspect is
`as described above the provision of small scale non
`traumatic test pieces as an external test electrode strip
`capable of using the naturally-arising small blood droplet
`from a needle-prick tester, Without tissue massage.
`Examples are described in more detail beloW.
`These small-scale electrodes are intended as single-use
`throWaWay articles and are utiliZed in conjunction With
`electrical circuitry and a readout means, to Which they must
`be easily attachable and detachable. Such circuitry and
`readout means is itself preferably embodied on a very small
`scale.
`We have accordingly found that the totality of the equip
`ment is subject to certain design constraints.
`Thus, it is further object of the invention in this form that
`the device should be nontraumatic to the user either physi
`cally e.g., if used With its oWn invasive probe or psycho
`logically by virtue of its appearance.
`It is further object of the invention in this form that the
`device should be capable, despite the small siZe of the
`throWaWay electrode and of the permanent circuitry/readout
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`components, or easy assembly and disassembly even by
`juvenile or elderly lay users.
`It is still a further object of the invention in this form to
`ensure the relatively expensive permanent circuitry/readout
`components should, despite their small siZe, be of a form
`Which minimiZes loss or damage.
`It is still a further object of the invention in this form to
`provide a device the display readings of Which are visible
`and understandable to a non-expert user.
`We have noW found that these and other objects of the
`invention can be met by assembly the circuitry/readout
`components into a housing resembling a pen/digital-Watch.
`According therefore another aspect of the present inven
`tion there is provided an assembly of circuitry and display
`means for use in producing a readout value as a diagnostic
`aid in human or veterinary medicine, housed in a pen-like
`holloW elongate housing having (a) at one end an electrically
`conductive socket suitable to receive the outer end of at least
`one detachable test member capable of producing an elec
`trical signal correlating With a physiological parameter to
`Which the test member is selectively sensitive and (b)
`toWards the other end a digital readout WindoW for exhib
`iting a numerical value corresponding to the parameter. A
`thermistor may also be used for temperature compensation.
`The person skilled in the art of designing medical equip
`ment Will appreciate that the invention extends not only to
`the pen-like assembly as de?ned above but also to the
`combination of such an assembly With an attached test
`member, and to the combination as a kit of interrelated parts
`of such an assembly With a plurality of test members suitable
`for one-off use.
`The term “pen-like” is a general limitation on siZe and
`shape. In functional terms, its characteristics are such that it
`can be held near the socket betWeen the thumb and the
`nearer one or tWo opposed ?ngers, With the elongate body
`resting on and extending beyond the fore?nger, but not to an
`extent that prejudices ?ne control of the socket end by the
`thumb and ?ngers. In numerical terms it can be from 10 to
`30 cm. long and from 0.5 to 3 cms across its maximum
`transverse dimension; more usually it Will be from 12 to 20
`cms. long and from 0.8 to 1.5 cms. across. It can be generally
`circular, or polygonal, in cross-section. Each detachable test
`member is usually a small-scale enZyme-coated sensor
`electrode, of the type discussed in US. Ser. No. 607,699,
`and especially such an electrode Where the enZyme is
`speci?cally glucose-catalyZing Whereby diabetic conditions
`can be measured. It may alternatively be a ?at external strip
`electrode dimensioned to operate on a small, non-expressed,
`blood droplet. The socket arrangement Will vary accord
`ingly.
`In one embodiment of the present invention, tWo or more
`sensor electrodes may be incorporated into a single test
`member. Again, the socket arrangement Will vary accord
`ingly.
`The readout means Will typically be a conventional seven
`segment display WindoW toWards the rearWard end of the
`“pen” as in conventional pen/Watches. In the case of the
`multiple sensor embodiment described in the preceding
`paragraph the display may be sWitchable betWeen each
`sensor’s discrete monitoring circuit, both the display and a
`single monitoring circuit may be sWitchable betWeen
`sensors, or a speci?c display may be provided for each of the
`sensors present.
`I. Constructional Features
`(a) Membrane Cover for Electrode
`Although the enZyme electrode should be in electrical
`contact With the liquid, it may be found valuable to exclude
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`7
`the sensor from interfering contact With larger molecules or
`tissue ?uid components. This can be done by a covering or
`surrounding membrane, depending on electrode geometry.
`Heat-shrinkable thin polymer tubing can be used as, or in
`connection With, such membranes.
`The membranes can be polymeriZed in situ. A particular
`valuable membrane is formed by polycarbonate, especially
`those polycarbonates sold under the Trade Marks “NUCLE
`OPORE” or “STERILIN”. When tissue ?uids are examined
`they may contain ascorbate; polycarbonate membranes do
`not permit the passage of ascorbate and thus virtually
`eliminate interference from that substance. Alternatively, a
`polyurethane membrane may be employed.
`(b) Type of Carbon
`Carbon foil, as strips, or carbon attached to metal meshes,
`of pyrolytic grade and knoWn by the Trade Marks “GRA
`PHOIL” and “PAPYEX” are much preferred for carbon
`ferrocene electrodes for use With glucose oxidase. Oxygen
`interference is minimal, there being less than 4% change in
`signal betWeen anaerobic and fully aerobic samples. Their
`physical nature is also very convenient for fabrication,
`especially of small-scale devices.
`II. Operational Features
`(a) Operational Potential
`Preferably operation should take place at a potential
`equivalent to +50 to +200 mV vs. SCE since intereference
`caused by oxidation of other chemical species present is
`thereby reduced.
`(b) Concentration Range
`Glucose oxidase can be used to monitor glucose concen
`trations of 0 to 40 mM, and glucose dehydrogenase at 0 to
`20 mM When immobilised on a carbon-ferrocene electrode.
`The sensor response is linear up to about 40 mM.
`(c) Response Times
`The glucose oxidase sensor Without membrane is kineti
`cally limited giving rapid response times i.e., about 20
`seconds to 95% of the steady-state current response.
`(d) Oxygen-Sensitivity
`Glucose dehydrogenase/ferrocene electrodes are totally
`oxygen-insensitive.
`(e) Use of Third Electrode
`In practice, a realistic device can achieve good perfor
`mance Without a third electrode, using Ag/AgCl as a refer
`ence counterelectrode, as described more fully beloW.
`(f) pH and Temperature
`Glucose oxidase electrodes shoW no change in current
`output betWeen pH6 and pH9, and are thus relatively
`pH-insensitive. They are temperature-stable up to 40° C. If
`necessary temperature compensation can be effected using a
`thermistor, or a constant temperature jacket may be used.
`Also, operating With the electrodes diffusion-limited mini
`mises temperature effects.
`(g) Storage of Electrodes
`Electrodes may be stored moist. Extended storage, over
`months or years, may be achieved by freeZe-drying or
`air-drying.
`Although the invention as de?ned above has been dis
`cussed in terms of the equipment used, it Will be appreciated
`that other aspects of the invention also present themselves.
`The totality of the equipment may include a replaceable or
`throWaWay cell; thus, the cell per se as de?ned above is an
`aspect of the invention, as are the combination of electrodes
`irrespective of details of cell design, and the individual
`electrodes, of novel con?guration. Methods of detecting the
`presence of, measuring the amount of or monitoring the
`level of one or more desired components (e.g., glucose) of
`a liquid mixture (e.g., tissue ?uid or liquids derived
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`therefrom) utiliZing the equipment cells or electrodes cells
`de?ned above are also a feature of the present invention.
`Finally, the present invention is concerned With the elec
`trical circuitry for operating the equipment as described.
`According to this aspect of the present invention there is
`provided a measuring device for use With an electron
`transfer electrode, comprising means for comparing an elec
`trical output of the electrode With an electronic reference and
`means for providing a signal related to the electrical output
`of the electrode.
`By employing an electronic reference rather than a cell of
`reference electrode a measurement using a sensor including
`an electron-transfer electrode may be made Without the use
`of a separate electrode as a reference.
`In a preferred embodiment of the invention, the electron
`transfer electrode is poised at a ?xed potential against a
`reference electrode, and the current ?oWing in the electron
`transfer electrode is measured.
`In the folloWing description of FIGS. 1 and 2 dimensions,
`materials, amount and proportions are given by Way of
`example only.
`In FIGS. 1 and 2, a strip of epoxy glass 1, 9.5><40><1.6 mm,
`has tWo 1 mm diameter holes 2 and 3 therein. A 9x9 mm
`piece of graphite tape of foil 4 is glued on one face, near the
`end to cover hole 2 and a 4x9 mm strip of silver foil 5 is
`glued adjacent thereto over hole 3. Wires 6 and 7 (FIG. 2)
`on the back enter holes 2 and 3 respectively for electrical
`connection With the respective electrode material 4 and 5,
`being glued in the holes by conductive epoxy resin 8. A
`stabiliZing layer of epoxy resin is present over at least part
`of the back e.g., a