CN100415896C - Creatine-kinase biosensor and its detergent preparation method - Google Patents
Creatine-kinase biosensor and its detergent preparation method Download PDFInfo
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- CN100415896C CN100415896C CNB2004100585134A CN200410058513A CN100415896C CN 100415896 C CN100415896 C CN 100415896C CN B2004100585134 A CNB2004100585134 A CN B2004100585134A CN 200410058513 A CN200410058513 A CN 200410058513A CN 100415896 C CN100415896 C CN 100415896C
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Abstract
The present invention discloses a creatine kinase biosensor and a preparation method of the reagent thereof, which belongs to the technical field of biosensors. The creatine kinase biosensor can quickly detect the concentration of total creatine kinase. The creatine kinase biosensor of the present invention is provided with a nanometer material layer which is arranged on the surfaces of two electrode of a capillarity channel, and a reaction reagent is fixed on the upper surface of the nanometer material layer, wherein the nanometer material layer is one kind of platinum black particles or high molecular nanometer material; the electrode surface with high activity and the nanometer material layer with high specific surface area are obtained. The preparation method of the reagent of the creatine kinase biosensor has the steps that two kinds of solutions are firstly prepared; then, the solutions are mixed with an electron acceptor solution to form the reagent. When an analyte which is a sample containing creatine kinase is dropped on a reaction region of a test bar through capillary action, the concentration of the total creatine kinase can be quickly detected by a test system of an electrochemical biosensor.
Description
Technical field
The present invention relates to the biosensor technology field, is the manufacture method of total creatine kinase strip of a kind of amperometric biosensor rapid detection and agents useful for same, the concentration that this strip can the total creatine kinase of rapid detection.
Technical background
Creatine kinase (Creatine Kinase) is present in the cytoplasm and plastosome of the muscle tissue of whole body and cerebral tissue, makes the regeneration with intracellular energy metabolism, Muscle contraction, ATP that the important kinases of direct relation be arranged.The reversibly following reaction of catalysis:
Annotate: CK: creatine kinase, Creatinephosphate: phosphocreatine, ADP: adenosine diphosphate (ADP), Creatine: creatine, ATP: Triphosaden, Mg: the magnesium ion that the creatine kinase reactive behavior is essential.
The mensuration of creatine kinase activity is one of the elementary item of the routine inspection of clinical diagnosis muscle disease, sacred disease, encephalopathic and heart disease.Cardiovascular disorder, especially myocardial infarction are major causes of death in the cardiovascular disease.Patient fall ill urgency, death risk height, creatine kinase increases in the several hrs after myocardial infarction, so quick diagnosis patient's creatine kinase is very necessary.In addition, prostate cancer, lung cancer, neural blastoma patient's creatine kinase level is higher than the normal people.And diabetics's hyperglycemia suppresses the expression of kinase activity in the muscle tissue, and it is the result of diabetes development or the reason that the diabetes state of an illness further develops that creatine kinase activity reduces.And swashing sour creatase detection is one of important indicator of sportsmen's athletic training intensity.Under the normal circumstances, muscle cell membrane structural integrity, function are normal, and creatine kinase (CK) seldom appears cytolemma.The micro-damage that training and training cause can make creatine kinase activity increase in the serum.Serum creatine kinase is the important sensitive indicator that evaluation muscle bears stimulation and the fine damage of skeletal muscle and adaptation and recovery.
At present, the method for mensuration serum creatine kinase commonly used is enzyme coupling connection test kit method, dry type biochemical analysis method and immunoassay, and this several method all needs expensive supporting detecting instrument.Instrument itself does not have the design function that stands ready and need for emergency treatment.Detecting instrument is subjected to the interference of other materials in the serum easily owing to adopt optics colorimetry principle etc., and detecting has error, and can not detect whole blood.
Summary of the invention
The objective of the invention is to go out to train application such as detection at myocardial infarction emergency case and sportsmen, provide a kind of fast, accurately, convenient, and can with the supporting portable biometric transmitter creatine kinase strip that can be used for whole blood test of portable small-sized instrument.
For achieving the above object, technical solution of the present invention provides a kind of creatine kinase bio-sensing device, comprises following structure:
At the insulativity upper surface of base plate pair of parallel electrode and contact conductor are arranged;
Two electrodes are respectively working electrode and counter electrode, and two electrodes top is covered with insulation layer;
Insulation layer laterally has a groove;
The groove top is coated with encapsulated layer;
The wicking action raceway groove is formed in the space that insulativity substrate, groove and encapsulated layer surround, wicking action raceway groove two ends are the capillary injection port, this wicking action raceway groove is the electrode reaction district, its two electrode surfaces at the wicking action raceway groove have a layer of nanomaterial, and the layer of nanomaterial upper surface is fixed with reaction reagent layer; Layer of nanomaterial, a kind of in platinum black particle or the high molecule nano material; To obtain the layer of nanomaterial of highly active electrode surface and high-specific surface area.
Described creatine kinase bio-sensing device, its described reaction reagent comprises electron acceptor, coupled reaction composite reagent, damping fluid, zymoexciter, enzyme reductive agent, antithrombotics and tensio-active agent.
Described creatine kinase bio-sensing device, its described high molecule nano material is the polymer Xylo-Mucine, wherein the polymer carboxymethylcellulosodium sodium content is 0.1～1%.
Described creatine kinase bio-sensing device, its described coupled reaction composite reagent reacts with assay, can produce the corresponding electric current of concentration with the total creatine kinase of assay;
In the group that electron acceptor is ferricyanate, methylene blue, ferrocene and derivative thereof, form stupid quinone, phenazine methosulfate, indophenols and derivative thereof and β-Nai Kun-4-Huang Suanjia any;
Damping fluid is any in the group formed of phosphoric acid buffer, TRIS damping fluid, MES damping fluid;
Zymoexciter is magnesium acetate or magnesium chloride;
The enzyme reductive agent is N-acetylcystein, reduced glutathion or contains a kind of of thiol compound;
Antithrombotics is any among heparin, oxalate, Citrate trianion and the EDTA;
Tensio-active agent is any in TritonX-100, diglycollic amide, alkylol amide phosphide salt, alcohol ether carboxylate, monoalkyl phosphoric acid esters, sodium laurylsulfonate or the alkylphenol polyoxyethylene.
Described creatine kinase bio-sensing device, its described coupled reaction composite reagent comprises: phosphocreatine, glucose, single adenosine phosphate (AMP), adenosine diphosphate (ADP) (ADP), coenzyme II (NADP) or nadide (NAD), hexokinase or glucokinase, glucose-6-phosphate dehydrogenase (G6PD), these several reagent of diaphorase all or part of.
A kind of preparation method of creatine kinase bio-sensing device agents useful for same comprises the following steps: a) at first phosphocreatine, glucose to be mixed with into solution R1; B) all or part of, damping fluid, zymoexciter, enzyme reductive agent, the antithrombotics with single adenosine phosphate (AMP), adenosine diphosphate (ADP) (ADP), coenzyme II (NADP) or nadide (NAD), hexokinase or glucokinase, glucose-6-phosphate dehydrogenase (G6PD) and these several reagent of diaphorase is mixed with into solution R2 with tensio-active agent; C) after the counter electrode surface activation process, solution R1, R2 and electron acceptor solution are mixed, drip on the electrode layer of nanomaterial uniformly then; D) to c) after the step, gained work in-process carried out drying, cover into encapsulated layer with insulating material; E) with slitting shear machine the strip array is cut into single strip, Vacuum Package is standby then.
Described preparation method of reagent thereof, its described c) counter electrode surface activation process in the step is with plasma method fast activating electrode surface.
Described preparation method of reagent thereof, its described d) step is c) to be placed on temperature be that drying covered encapsulated layer with insulating material after 20～25 minutes in 35～37 ℃ the loft drier to step gained work in-process.
Described creatine kinase bio-sensing device, its detection system adopt electric current to detect.
Product of the present invention relatively has many advantages with at present domestic and international product.The method of strip of the present invention and introduction is compatible with the portable type current type instrument of business-like series, therefore can carry, detect simple and fast; Because the production technique of disposable test strips adopts advanced MEMS (micro electro mechanical system) (MEMS) technology, both guaranteed the reliability and the consistence that detect, reduced the production cost of strip again; Adopt the fixing means of simple and effective enzyme, the principle that has adopted electrochemical source of current to detect, accurate than the colorimetry principle, make that the production of strip is more easy; Strip adopts the capillary design, makes sample introduction faster, need blood volume still less (3 μ l), and the response of system is fast, and is highly sensitive.
Description of drawings
Fig. 1 is the layering synoptic diagram of two electrode systems that design of the present invention;
Fig. 2 electrode plane figure of the present invention.
The cyclic voltammetric comparison diagram of the different concns polymer carboxymethyl cellulose (CMC) of Fig. 3 embodiment of the invention.
The creatine kinase real-time response figure of Fig. 4 embodiment of the invention.
The creatine kinase linear relationship chart of Fig. 5 embodiment of the invention.
Embodiment:
The preparation method of transmitter test strip electrode of the present invention is with reference to patent applied for: 00410030214.X and 200410031948.X.
With the technology of plasma method fast activating electrode surface with reference to patent applied for 02154524.3.
The invention provides a kind of one deck high molecular nanometer particulate method of in the electrode surface reaction zone, assembling.By evenly drip the wetting ability macromole Xylo-Mucine of different concns at electrode surface, after drying under 37 ℃ of temperature condition, obtain to have the electrode nanometer layer in different apertures.This method has further been strengthened the wetting ability of electrode surface, has increased the effective surface area of electrode.Not only help the immobilization of enzyme reagent, and increased the electrochemical activity of electrode.
The invention provides fixation reaction compositions and methods on nano particle.
The present invention is fixed on reaction reagent and forms reagent layer on the electrode, the reaction reagent that is fixed on the high molecular nanometer layer comprises electron acceptor, react with assay and can produce composite reagent with the coupled reaction of the corresponding electric current of analyte concentration, also comprise damping fluid, zymoexciter, reductive agent, antithrombotics and tensio-active agent simultaneously.
The reaction principle of agent combination of the present invention institute foundation is as follows:
Creatine phosphate phosphocreatine wherein; The Creatine creatine; The CK creatine kinase; Glucose glucose; Glucose-6-phosphate 6-phosphoric acid-glucose; Gluconolactone-6-phosphate6-phosphoglucose lactone; The HK hexokinase; The GPD glucose phosphate dehydrogenase; NADPH NADP
+(coenzyme II) reduced form/NADP+; The DIA diaphorase; Fe (CN)
6 4-/ Fe (CN)
6 4-Reduced form/oxidized form the Tripotassium iron hexacyanide
At first phosphocreatine, the glucose with proper concn is mixed with into solution R1, single adenosine phosphate (AMP), adenosine diphosphate (ADP) (ADP), coenzyme II (NADP), hexokinase, glucose-6-phosphate dehydrogenase (G6PD), diaphorase, magnesium acetate, reduced glutathion, EDTA, the TritonX-100 of proper concn are mixed with into solution R2, after the electrode surface activation treatment is good, again solution R1, R2 and potassium ferricyanate solution are mixed, evenly drip on electrode high molecular nanometer layer with point sample instrument then.In 35～37 ℃ of loft drier, after dry 20～25 minutes, cover encapsulation, the strip array is cut into single strip with slitting shear machine with insulating material.Vacuum Package then.Standby.
The suitable concentration range 10～1000mM of phosphocreatine in the reagent, it optimizes reaction range: 50mM～100mM.The suitable concentration range 10～1000mM of glucose, it optimizes reaction range: 50mM～100mM.The suitable concentration range 1～100mM of single adenosine phosphate, it optimizes reaction range: 5mM～20mM.The suitable concentration range 0.5～100mM of adenosine diphosphate (ADP), it optimizes reaction range: 1mM～20mM.Coenzyme II suitable concentration range 0.5～100mM, it optimizes reaction range: 1mM～20mM.Hexokinase, glucose-6-phosphate dehydrogenase (G6PD), diaphorase are suitable for concentration range 1～2000U/ml, and it optimizes reaction range: 1～40U/ml.
The electron acceptor that plays redoxomorphism is a kind of by what select the group of forming from ferricyanate, methylene blue, ferrocene and derivative thereof, to stupid quinone, phenazine methosulfate, indophenols and derivative thereof and β-Nai Kun-4-Huang Suanjia; By electron mediator the electronics that produces in the enzyme reaction process is transferred to electrode surface from the enzyme reaction center, make the response speed of current mode Enzyme sensor and detection sensitivity be improved, reduced the voltage of reaction simultaneously.Described electron acceptor is a potassium ferricyanate, makes operating potential reduce to 0.2V, reduces the interference of other active substance in the blood.The suitable concentration range of potassium ferricyanate: 0.1mM～300mM, it optimizes reaction range: 10mM～100mM.
Damping fluid is select in the group formed of phosphoric acid buffer, TRIS damping fluid, MES damping fluid a kind of; Described damping fluid is a phosphoric acid buffer.Damping fluid is used to provide the reaction environment of an acidity-basicity ph value stabilization, and the best acidity-basicity ph of reaction is 6～8, concentration range: 10～100mM.
In the creatine kinase reaction, magnesium ion is necessary, and zymoexciter is magnesium acetate or magnesium chloride.Select magnesium acetate for use, its suitable concentration range: 1～100mM, optimum concentration range: 5～20mM.
Because most of creatine kinases exist with inactive form in the serum, measure its activity for accurate quantification, it must be activated, enzyme reductive agent commonly used is that N-acetylcystein, reduced glutathion etc. contain a kind of of thiol compound; Described enzyme reductive agent is a reductive glutathione.Because sulfhydryl compound poor stability in solution, therefore, reductive glutathione need be used freshly prepared solution.Its suitable concentration range: 0.1～10mM, optimum concentration range: 0.1～5mM.
Because in the whole blood test process, blood is easy to solidify, general blood began to solidify at 2～5 minutes, therefore, carried out in order to react smoothly, needed to add in reagent antithrombotics.Antithrombotics is a kind of for what select from heparin, oxalate, Citrate trianion and EDTA; Described antithrombotics is EDTA.Its suitable concentration range: 0.1～10mM, optimum concentration range: 0.5～5mM.
Described tensio-active agent is TritonX-100.Add the tensio-active agent of 0.05～1% non-ionic type.Preferred 0.1～0.5% concentration tensio-active agent.
Above-mentioned various purposes of the present invention, method, characteristics and advantage are by obtaining more detailed explanation below in conjunction with drawings and Examples.
Fig. 1, Fig. 2 are the two electrode figure that the present invention designs.At first, adopt MEMS (micro electro mechanical system) (MEMS) technology to prepare film gold electrode 2,3 on insulativity base material 1, be provided with wicking action raceway groove 7 in sticking adding in second insulation layer, 4, the second insulation layers 4 on the film gold electrode 2,3, wicking action raceway groove 7 forms reaction zone.Then, electrode 2,3 is assembling one deck polymer Xylo-Mucine (CMC) nano-particle layer 8 in the wicking action raceway groove 7 at electrode 2,3 reaction zones after ion cleaning machine activation, 37 ℃ of electric drying oven with forced convection inner dryings 20 minutes, takes out standby.
Add reagent then, reagent is: phosphocreatine: 100mM; Glucose: 50mM; Magnesium acetate: 20mM; Single adenosine phosphate (AMP): 20mM; Adenosine diphosphate (ADP) (ADP): 5mM; Nicotine adenosine two phosphatase nucleic acids (NADP): 5mM; Glucose 6-phosphate dehydrogenase: 4U/ml; Hexokinase: 4U/ml; Diaphorase: 2U/ml; Gsh: 3mM; EDTA:1mM; TritonX-100:0.2%; Potassium ferricyanate: 25mM; Phosphoric acid buffer: 0.01M.
5 μ l mentioned reagent drips of solution are added in the wicking action raceway groove 7, are covered with on the nano-particle layer 8 of one deck polymer carboxymethyl cellulose (CMC), drying is 20 minutes in 37 ℃ loft drier, forms reagent layer 9.From loft drier, take out strip, add a cover transparent insulation encapsulated layer 5, form wicking action raceway groove 7, and the capillary injection port 10 at two ends.With slitting shear machine the strip array is cut into single strip.Vacuum Package then.
On the electrode 2,3 for preparing, after cleaning with ionized water, drip 0.5% and 1% polymer carboxymethyl cellulose (CMC), 37 ℃ of electric drying oven with forced convection inner dryings 20 minutes.After the taking-up, detect its current voltage characteristic.Among Fig. 3, on electrode 2,3, the polymer carboxymethyl cellulose (CMC) of assembling different concns: (1) 0%; (2) 0.5%; (3) 1% cyclic voltammogram.Scanning speed: 100mV/s, solution-based phase: 0.01M phosphoric acid buffer (pH7.4), 0.03MK
3Fe (CN)
6As seen from Figure 3, along with polymer carboxymethyl cellulose (CMC) concentration increases, its electric current increases, but increases with polymer carboxymethyl cellulose (CMC) concentration, and the high molecular nanometer film thickens, and electron transport speed reduces, and influences speed of response; The background response electric current increases.Therefore need to select suitable concentration.General selection 0.1～1% comes to determine as the case may be.On electrode 2,3 surfaces of assembling high molecular nanometer film, surface-area obviously increases, add aqueous reaction reagent after, layer of nanomaterial 8 forms sol layer with reagent, therefore produces bigger reaction current, augmenting response sensitivity.
Between the working electrode of strip and reference electrode, apply+voltage of 0.2V.On the electrode 2,3 of having modified macromolecular material Xylo-Mucine (CMC) nanometer layer 8, drip mix reagent, after question response electric current (background current) is steady, the creatine kinase damping fluid that adds different activities, have an effect with reagent and high molecular nanometer layer 8, produce electric current, flow out, can rapid detection on the electrochemica biological sensor detection system go out active corresponding electric current with total creatine kinase through contact conductor 6.The electric current that produces is directly proportional with creatine kinase activity.And in very wide scope, present good linear dependence.Therefore, this transmitter strip has excellent application value.See Fig. 4 a, for biosensor to different concns creatine kinase real-time response curve.Creatine kinase activity: (1) 800U/ml; (2) 400U/ml; (3) 80U/ml; (4) 8U/ml; (5) 800U/L; (6) 400U/L; (7) 80U/L; (8) 40U/L; (9) 8U/L.Fig. 4 b is Fig. 4 a dotted portion enlarged view.Fig. 5 is the creatine kinase linear relationship chart.Fig. 5 a is a creatine kinase high reactivity scope, the reaction times: 140 seconds; Fig. 5 b is a field of activity at the bottom of the creatine kinase, the reaction times: 280 seconds.
The present invention has used preferred embodiment to describe, and preferred embodiment is for illustrative purposes, rather than limitation of the present invention.The present invention can be many modifications and changes on the basis of the above description.Therefore, within the scope of the appended claims, the present invention can have other implementation that is not above-mentioned.For example: thus the agent prescription of other combination, on reagent layer, cover filtering membrane eliminate erythrocytic interference and be used for whole blood creatine kinase detection, can adopt optical detection, can adopt electrode reaction district that other nano particle and non-nano material modify, the preparation form of three electrode forms, thick membrane electrode etc.
Claims (9)
1. creatine kinase bio-sensing device comprises following structure:
At the insulativity upper surface of base plate pair of parallel electrode and contact conductor are arranged;
Two electrodes are respectively working electrode and counter electrode, and two electrodes top is covered with insulation layer;
Insulation layer laterally has a groove;
The groove top is coated with encapsulated layer;
The wicking action raceway groove is formed in the space that insulativity substrate, groove and encapsulated layer surround, and wicking action raceway groove two ends are the capillary injection port, and this wicking action raceway groove is the electrode reaction district, it is characterized in that:
Two electrode surfaces at the wicking action raceway groove have a layer of nanomaterial, and the layer of nanomaterial upper surface is fixed with reaction reagent layer; Layer of nanomaterial, a kind of in platinum black particle or the high molecule nano material; To obtain the layer of nanomaterial of highly active electrode surface and high-specific surface area.
2. creatine kinase bio-sensing device as claimed in claim 1 is characterized in that: described reaction reagent comprises electron acceptor, coupled reaction composite reagent, damping fluid, zymoexciter, enzyme reductive agent, antithrombotics and tensio-active agent.
3. creatine kinase bio-sensing device as claimed in claim 2 is characterized in that: described coupled reaction composite reagent, react with assay, and can produce the corresponding electric current of concentration with the total creatine kinase of assay;
In the group that electron acceptor is ferricyanate, methylene blue, ferrocene, form stupid quinone, phenazine methosulfate, indophenols and β-Nai Kun-4-Huang Suanjia any;
Damping fluid is any in the group formed of phosphoric acid buffer, TRIS damping fluid, MES damping fluid;
Zymoexciter is magnesium acetate or magnesium chloride;
The enzyme reductive agent is N-acetylcystein, reduced glutathion or contains a kind of of thiol compound;
Antithrombotics is any among heparin, oxalate, Citrate trianion and the EDTA;
Tensio-active agent is any in TritonX-100, diglycollic amide, alkylol amide phosphide salt, alcohol ether carboxylate, monoalkyl phosphoric acid esters, sodium laurylsulfonate or the alkylphenol polyoxyethylene.
4. creatine kinase bio-sensing device as claimed in claim 3 is characterized in that: described coupled reaction composite reagent comprises: phosphocreatine, glucose, single adenosine phosphate AMP, adenosine diphosphate (ADP) ADP, coenzyme II NADP or nadide NAD, hexokinase or glucokinase, glucose-6-phosphate dehydrogenase (G6PD), these several reagent of diaphorase all or part of.
5. creatine kinase bio-sensing device as claimed in claim 3, it is characterized in that the concentration range of described electron acceptor: 10mM～100mM, buffer concentration scope: 10～100mM, zymoexciter concentration range: 0.1～10mM, enzyme reductant concentration scope: 5～20mM, antithrombotics concentration range: 0.5～5mM, surfactant concentration ranges: 0.1～0.5%.
6. creatine kinase bio-sensing device as claimed in claim 4 is characterized in that: in the described coupled reaction composite reagent, the concentration range of phosphocreatine is 10～1000mM; The concentration range of glucose is 10～1000mM; The concentration range of single adenosine phosphate is 1～100mM; The concentration range of adenosine diphosphate (ADP) is 0.5～100mM; Coenzyme II concentration range is 0.5～100mM; The concentration range of hexokinase, glucose-6-phosphate dehydrogenase (G6PD), diaphorase is 1～2000U/ml.
7. the preparation method of a creatine kinase bio-sensing device is characterized in that: comprise the following steps: a) at first phosphocreatine, glucose to be mixed with into solution R1; B) all or part of, damping fluid, zymoexciter, enzyme reductive agent, the antithrombotics with single adenosine phosphate AMP, adenosine diphosphate (ADP) ADP, coenzyme II NADP or nadide NAD, hexokinase or glucokinase, glucose-6-phosphate dehydrogenase (G6PD) and these several reagent of diaphorase is mixed with into solution R2 with tensio-active agent; C) after the counter electrode surface activation process, solution R1, R2 and electron acceptor solution are mixed, drip on the electrode layer of nanomaterial uniformly then; D) to c) after the step, gained work in-process carried out drying, cover into encapsulated layer with insulating material; E) with slitting shear machine the strip array is cut into single strip, Vacuum Package is standby then.
8. the preparation method of creatine kinase bio-sensing device as claimed in claim 7 is characterized in that: counter electrode surface activation process in the step described c) is with plasma method fast activating electrode surface.
9. the preparation method of creatine kinase bio-sensing device as claimed in claim 7 is characterized in that: the step described d) is c) to be placed on temperature be that drying covered encapsulated layer with insulating material after 20～25 minutes in 35～37 ℃ the loft drier to step gained work in-process.
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| JP4723311B2 (en) * | 2005-08-17 | 2011-07-13 | シスメックス株式会社 | Reagent for measuring creatine kinase activity |
| CN104485232B (en) * | 2014-12-18 | 2017-06-06 | 中国科学院上海硅酸盐研究所 | A kind of preparation method of used by dye sensitization solar battery to electrode |
Citations (4)
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|---|---|---|---|---|
| WO1996012188A1 (en) * | 1994-10-18 | 1996-04-25 | Surface Active Limited | Diagnostic method and apparatus for performing the method |
| WO1997037218A2 (en) * | 1996-03-29 | 1997-10-09 | Byk Gulden Italia S.P.A. | Automatic diagnostic apparatus |
| JP2775055B2 (en) * | 1989-02-08 | 1998-07-09 | 新日本無線株式会社 | Biosensor |
| CN1502988A (en) * | 2002-11-27 | 2004-06-09 | 浙江贝特生物园区发展有限公司 | Nano silicon membrane biological sensor |
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2004
- 2004-08-16 CN CNB2004100585134A patent/CN100415896C/en not_active Expired - Fee Related
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2775055B2 (en) * | 1989-02-08 | 1998-07-09 | 新日本無線株式会社 | Biosensor |
| WO1996012188A1 (en) * | 1994-10-18 | 1996-04-25 | Surface Active Limited | Diagnostic method and apparatus for performing the method |
| WO1997037218A2 (en) * | 1996-03-29 | 1997-10-09 | Byk Gulden Italia S.P.A. | Automatic diagnostic apparatus |
| CN1502988A (en) * | 2002-11-27 | 2004-06-09 | 浙江贝特生物园区发展有限公司 | Nano silicon membrane biological sensor |
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| 纳米材料与微电极生物传感器. 王利，等..微纳电子技术，第7/8期. 2003 |
| 纳米材料与微电极生物传感器. 王利，等..微纳电子技术，第7/8期. 2003 * |
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