CN103105426A - Urine glucose testing method and biosensor used in method - Google Patents
Urine glucose testing method and biosensor used in method Download PDFInfo
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- CN103105426A CN103105426A CN2013100183366A CN201310018336A CN103105426A CN 103105426 A CN103105426 A CN 103105426A CN 2013100183366 A CN2013100183366 A CN 2013100183366A CN 201310018336 A CN201310018336 A CN 201310018336A CN 103105426 A CN103105426 A CN 103105426A
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Abstract
The invention aims to provide a urine glucose testing method through which the urea conductivity is tested at high precision and high reliability and full and accurate correction can be performed as well as a biosensor used in the method. The biosensor is easy in manufacturing process, low in cost and convenient to use. The method comprises the following steps: a, performing oxidation reduction on glucose in urea in the presence of an amboceptor by utilizing an oxidation-reduction enzyme; b, detecting the generated oxidation current or reduction current at the moment through an electrode system, and calculating the glucose concentration based on the current value; and c, testing the conductivity in the urea, and correcting the glucose concentration in the urea by utilizing the conductivity. The biosensor comprises an insulating substrate, an electrode system positioned on the insulating substrate, an electric insulating layer and a reaction layer which is positioned in an electrochemical electrode reaction region.
Description
Technical field
The present invention relates to field of biological detection, the biology sensor that specifically uses in the assay method of glucose in urine and the method.
Background technology
In order to control metabolic situation, particularly for diabetes cases, need the concentration of glucose in control volume circulation termly.This is not only necessary for the type i diabetes patient who wants insulin injection, is also that so the latter accounts for 85% of this glycometabolism disease for the type ii diabetes patient.It is expected that, to world diabetic number in 2010 will reach 200,012,100 (Hauner, H; Dtsch Med Wochenschr (German medical science weekly) 1998; 123; 777-82; Pharmaco Economlcs (medication economics) 1995; 8 (Suppl, I) 28-71; ).
In view of the high incidence of diabetes, only there is the conventional blood glucose measurement that is undertaken by skin penetrating can not satisfy needs and the hope that regular metabolism is controlled.This needs the patient to overcome the mental handicape of stabbing skin before each the measurement.For Most patients, do not need to measure blood sugar, and as long as controlled the concentration of glucose in urine.If surpassed the threshold value of kidney by metabolic stimulation, just cause the outer glucose of physiological requirements to be discharged through kidney as glucose in urine, the glucose in urine just is presented as can detected concentration.Therefore, although blood sugar test is quite universal, glucose in urine detects as a kind of very easy noninvasive detection method of operation of using, and still deeply is subjected to considerable part diabetic's welcome.
A kind of method of glucose in urine of measuring makes light polarization and dextrorotation based on the characteristic of glucose.Said polarization photometer has just utilized this specific character, wherein light is the estimating of actual concentration of glucose (Poege, A.W.Z med.Labortechnik (medical experimental chamber diagnostic techniques) 17 (1976) 59-79) from the angle of plane of polarization deflection.
Usually adopt the glucose in urine Test paper for the screening family of prevention use and the method for the glucose content in a human urine, this class test paper majority is to produce the color reaction of glucose compliance on the zymochemistry basis.The shortcoming of these test strips is poor accuracy, the half-quantitative detection that can only be used for glucose in urine, and remolding sensitivity is lower, generally all reach 50mg/dl at glucose in urine content, even 100mg/dl just can demonstrate the positive, also must be with accurate judged result of strict control time of clock during test, thus measurement result is very unreliable.And only have the people that can differentiate color to adopt, and in diabetic's, particularly type ii diabetes gerontal patient, considerable people is colour blindness, debates at least look inaccurate.
Above-mentioned polarization photometer is the clinical chemistry that is used for that is equipped with as laboratory equipment, and the glucose of expressing reliably in daily metabolism control in urine exists and the conventional method of concentration and Tes-Tape is not suitable for being used as.Therefore research and development more and more concentrates on enzymatic-electrochemical measuring device and the application aspect glucose assays thereof.2008, a company development of Japan has been released and a kind ofly can have been carried out the biology sensor glucose in urine tester that quantitatively detects to glucose in urine, but because it adopts fixed biologically sensor detection head, expensive, use cost is high, and detection head is repeatedly Reusability, needs to clean, and also can face some problems of sterilization aspect.
In urine, measurement causes serious interference to various impurity to glucose in urine, no matter be colorimetric reflectometry or electrochemica biological sensor method.Exist the amount of chaff interference to have than big difference in the different specific weight urine, contain the reductibility chaff interferences such as more Vc, uric acid in baruria, the a large amount of electrolyte ions that contain in baruria, metallic ion all can affect the specificity that glucose in urine detects, and cause test result inaccurate.
One is waiting for ratification, is being called the patent of " the electric pole type Tes-Tape with sample pre-treatments function ", the problem of disturbing in order to solve different urines, application of sample section at the electric pole type Tes-Tape adds the film that has soaked treating agent, to eliminating to disturb, effect is preferably arranged.But, due to the needs pre-treatment film, the strict control of the production and processing of pre-treatment film, the precision of guarantee Tes-Tape, even strict control sometimes, the test paper precision of production still can be relatively poor; In addition, due to the strict production control of needs, increased efficient and the cost of processing.
Existing glucose in urine method of testing, otherwise sensitivity is on the low side, affected greatly by various chaff interferences, causes test not accurate enough; Manufacture craft is numerous and diverse, and cost is high, uses inconvenient not.
For convenient, the painless analysis insulin state of diabetes patient and non-diabetic people, need that a kind of measurement sensitivity is high, accurate, the method for glucose in reliable test urine.The glucose in urine test is more accurate, reliable, and the elimination of interference, test precision etc. also have area for improvement.
Summary of the invention
Therefore, the object of the present invention is to provide by measuring the urine conductivity with high precision and high reliability can be fully and the bio-sensing that uses in the urinary glucose determination method of correctly proofreading and correct and the method, and manufacture craft is simple, and cost is low, and is easy to use.
In order to reach above-mentioned purpose, assay method of the present invention is the assay method of the glucose in urine, steps of the method are:
A under the existence of amboceptor, utilizes oxidoreducing enzyme that the glucose oxidase in urine is reduced;
B detects by electrode system oxidation current or the reduction current that produce this moment, take above-mentioned current value as the basic calculation concentration of glucose;
C measures the conductivity in above-mentioned urine and utilizes this conductivity to proofread and correct the correcting process of urine glucose concentration; The assay method of conductivity is: prepare to contain working electrode and to the electrode system of electrode, do not configure amboceptor on the working electrode in above-mentioned two electrodes, and configure amboceptor on to electrode; Urine is directed into above-mentioned electrode system, under this state, above-mentioned electrode system is applied voltage, detect thus and flow to above-mentioned two interelectrode oxidation current or reduction currents, take this current value as the basic calculation conductivity value.
Said oxidoreducing enzyme herein, mean can with sample in subject matter to be measured produce the enzyme of redox reaction.It is different and different that the enzyme class of using is looked the subject matter of institute's wish detection.For example, when wish detected concentration of glucose, selected enzyme was glucose oxidase or glucose dehydrogenase.
Said electron mediator herein, mean can with the substance reaction that produces through aforementioned oxidoreducing enzyme effect after, itself can be reduced into reducing condition by the state of oxidation and get material.When electron mediator becomes reducing condition, can be by applying an impressed voltage on pole reagent paper, impelling electron mediator to be replied by the reducing condition reversed reaction is the state of oxidation, this moment chemical reaction current potential, resistance or curent change, the joint that can conduct to the electrode system other end by working electrode and the reference electrode of electrode system.When this joint is connected with a signal receiver, current potential, resistance or the curent change of aforementioned chemical reaction can be received, and signal is converted to by a display device concentration that detects subject matter.Electron transfer mediator is complex compound or the benzoquinone compound of the potassium ferricyanide, ferrocene and derivant thereof, ruthenium.According to a preferred embodiment of the present invention, this electron mediator can be the potassium ferricyanide.Preferably, the content of this electron mediator is that 3% (m/m) is to 20% (m/m).
Said damping fluid, bonding agent and surfactant herein mean the material that can help be attached to after oxidoreducing enzyme and electron mediator drying on insulating substrate, maybe can protect redox material.Described damping fluid is a kind of in the group that forms of phosphate buffer, MES damping fluid, citrate buffer solution; Described bonding agent adopts one or more in starch, dextrin, gelatin, hydroxyethyl cellulose, carboxymethyl cellulose, sodium alginate; Described surfactant is TritonX-100, TW-20.The method that the enzyme reaction film is fixed on electrode can be absorption method, cross-linking method, sol-gel process, adhesive method etc.The absorption method immobilized enzyme only need to directly drip enzyme liquid on electrode surface, and the hydrophobic grouping of enzyme and hydrophobic electrode surface adsorb through Van der Waals force; The cross-linking method immobilized enzyme can add the cross-linking reagents such as glutaraldehyde in enzyme liquid.The sol-gel process immobilized enzyme can incorporate enzyme in the reagent such as silica hydrosol, titania hydrosol, gelatin, forms gel after the electrode surface drying.The adhesive method immobilized enzyme can be made into printable enzyme slurry with enzyme and bonding agent and other auxiliary materials, and serigraphy is on electrode surface.Electron transfer mediator, damping fluid and surfactant can be mixed with enzyme and drip or be screen-printed to electrode surface.
Biology sensor of the present invention is used for by redox glucose and utilizes electrode detection to measure above-mentioned urine glucose by oxidation current or reduction current that this reaction produces, and it is divided into glucose in urine test section and enrichment factor test section:
The glucose in urine test section has the 1st electrode system and reagent layer, and the mentioned reagent layer has oxidoreducing enzyme and the amboceptor of glucose in urine.Under the existence of amboceptor, by above-mentioned oxidoreducing enzyme with the glucose in urine redox, the redox electric current when utilizing above-mentioned the 1st electrode detection to apply voltage, thus measure glucose in urine concentration.
The enrichment factor test section has working electrode and to electrode, does not configure amboceptor on above-mentioned working electrode, and configure amboceptor on to electrode; Above-mentioned urine is directed into above-mentioned electrode system, applies voltage, detect thus the current value that flows, thereby measure the enrichment factor of above-mentioned urine.Proofread and correct above-mentioned glucose in urine concentration by this enrichment factor.
Biology sensor of the present invention, it comprises:
One insulating substrate;
One is positioned at the electrode system on insulating substrate, and this electrode system is provided with three strip conductive electrodes, is respectively contrast electrode, working electrode, enrichment factor mensuration electrode;
One part is covered in the electric insulation layer on electrode system, makes the electrode system zone that exposes not covering electric insulation layer, forms one and holds electrochemical electrode reaction zone and the vent port in sample solution space;
One is positioned at the responding layer of electrochemical electrode reaction zone, and this responding layer comprises at least a oxidoreducing enzyme, plants electron mediator, also comprises damping fluid, bonding agent and surfactant; Responding layer only covers contrast electrode, working electrode, does not measure electrode and do not cover enrichment factor.
Said insulating substrate, mean to have straight surface and the thin-layer tablet of electrical insulation characteristics herein.Preferably, this insulating substrate is selected from the following group who forms: Polyvinylchloride (PVC) sheet, glass fiber sheets (FR-4), polyester, bakelite sheet, PET sheet, PC sheet, PP sheet, PE sheet, PA sheet, PS sheet, glass sheet and potsherd (CEM-I).According to of the present invention one better enforcement, this insulating substrate is PC or PET sheet.
Said electrode system herein means to comprise having at least three to separate and not contacted electrode mutually, forms respectively working electrode, reference electrode and enrichment factor and measures electrode.According to of the present invention one better enforcement, electrode system system covers so that electric insulation layer is local, make and be exposed to electrical isolation separation layer one end and form respectively working electrode, reference electrode, enrichment factor and measure electrode and be connected with the electrochemical electrode reaction zone, the other end forms the joint that working electrode, reference electrode, enrichment factor are measured electrode, and the pick-up unit that can produce with test sample electrical effect when the electrochemical reaction is connected.Preferably, the material of this electrode system can be selected the conductive slurry-like material that is fit to screen painting, such as but not limited to carbon paste, gold size, elargol, carbon silver epoxy glue, volatility graphite or copper glue or its combination (printing again carbon paste after for example first printing elargol with wire mark), or the conductive slurry-like material of other any suitable screen painting.
Said electrical isolation separation layer herein, the formed thin layer of material that means to have electrical insulation property.Preferably, the material of this electrical isolation separation layer can be selected electrical insulating property pulpous state material or the electrical insulating property adhesive tape that is fit to screen painting, the insulating tape of for example insullac of heated drying type or ultraviolet light drying type, or PVC or PET material.Preferably, to have thickness be 0.1 to 0.25 millimeter to this electric insulation layer.
According to of the present invention one better enforcement, this electrical isolation separation layer part is covered in on-chip electrode system, makes the electrode system zone that exposes not covering electric insulation layer, forms one and holds electrochemical electrode reaction zone and the vent port in sample solution space.
Said responding layer, mean to be positioned at above-mentioned electrochemical electrode reaction zone herein, contains the biological reagent with the measured matter reaction.Responding layer comprises enzyme and a kind of electron transfer mediator with the measured matter reaction, also contains simultaneously the composite thin layer of damping fluid, bonding agent and surfactant.Preferably, after the mode that this composite can apply or drip adds to this responding layer, dry under 60 ℃.
In the assay method and biology sensor of glucose in urine of the present invention, the above-mentioned working electrode of above-mentioned urine enrichment factor determination part and the voltage that applies between electrode is preferably greater than the voltage that equals water electrolysis, be preferably the scope of 1-5V, more preferably the scope of 1-3V.By applying the voltage more than or equal to water electrolysis voltage, can only depend on the concentrated electric current of urine with higher sensitivity determination, and can access the impact that is not subjected to existing other redox materials in urine, the stable electric current that does not rely on individual differences.In addition, application time is for example 1-10 second, 1-5 second more preferably.
Thus, the invention is characterized in the mensuration of the enrichment factor of urine.That is, when measuring the enrichment factor of urine, by only configuring amboceptor on to electrode, can detect simply with high precision and high reliability the electric current of the enrichment factor of reflection urine.Therefore, by assay method of the present invention, biology sensor and determinator, the enrichment factor of the urine of measuring take this high precision and high reliability is proofreaied and correct glucose in urine concentration as the basis, therefore can fully and correctly proofread and correct, glucose in urine concentration after consequently proofreading and correct is more high precision and high reliability also, manufacture craft is simple, and cost is low, and is easy to use.
Description of drawings
Accompanying drawing described herein is used to provide a further understanding of the present invention, consists of a part of the present invention, does not consist of limitation of the invention.In the accompanying drawings:
Fig. 1 is the structural representation of the first embodiment of the present invention;
Fig. 2 is the STRUCTURE DECOMPOSITION figure of embodiment shown in Figure 1;
Fig. 3 is the sensor of the expression embodiment 1 time dependent figure of response current value (nA) with respect to the applying of voltage, enrichment factor;
Fig. 4 is the sensor of the expression embodiment 1 time dependent figure of response current value (nA) with respect to the applying of voltage (1V), enrichment factor;
Fig. 5 is the sensor of the expression embodiment 1 time dependent figure of response current value (nA) with respect to the applying of voltage (2V), enrichment factor;
Fig. 6 is the sensor of the expression embodiment 1 time dependent figure of response current value (nA) with respect to the applying of voltage (3V), enrichment factor;
Fig. 7 is the sensor of the expression embodiment 1 time dependent figure of response current value (nA) with respect to the applying of voltage (4V), enrichment factor;
Fig. 8 is the sensor of the expression embodiment 1 time dependent figure of response current value (nA) with respect to the applying of voltage (5V), enrichment factor;
Number in the figure is:
1 substrate; 2 pairs of electrodes; 3 working electrodes; 4 enrichment factors are measured electrode; 5 dielectric isolation layers; 6 Process windows; 7 enzyme reaction films; The sample groove is led in 8 siphons.
Embodiment
The invention provides the biology sensor that uses in the assay method that correctly to proofread and correct thus glucose in urine by measuring the enrichment factor of urine with high precision and high reliability and the method.Biology sensor of the present invention, it comprises:
One insulating substrate
One is positioned at the electrode system on insulating substrate, and this electrode system is provided with three strip conductive electrodes, is respectively contrast electrode, working electrode, enrichment factor mensuration electrode;
The part is covered in the electric insulation layer on electrode system, makes the electrode system zone that exposes not covering electric insulation layer, forms one and holds electrochemical electrode reaction zone and the vent port in sample solution space;
One is positioned at the responding layer of electrochemical electrode reaction zone, and this responding layer comprises at least a oxidoreducing enzyme, plants electron mediator, also comprises damping fluid, bonding agent and surfactant; Responding layer only covers contrast electrode, working electrode, does not measure electrode and do not cover enrichment factor.
Said insulating substrate, mean to have straight surface and the thin-layer tablet of electrical insulation characteristics herein.Preferably, this insulating substrate is selected from the following group who forms: Polyvinylchloride (PVC) sheet, glass fiber sheets (FR-4), polyester, bakelite sheet, PET sheet, PC sheet, PP sheet, PE sheet, PA sheet, PS sheet, glass sheet and potsherd (CEM-I).According to of the present invention one better enforcement, this insulating substrate is PC or PET sheet.
Said electrode system herein means to comprise having at least three to separate and not contacted electrode mutually, forms respectively working electrode, reference electrode and enrichment factor and measures electrode.According to of the present invention one better enforcement, electrode system system covers so that electric insulation layer is local, make and be exposed to electrical isolation separation layer one end and form respectively working electrode, reference electrode, enrichment factor and measure electrode and be connected with the electrochemical electrode reaction zone, the other end forms the joint that working electrode, reference electrode, enrichment factor are measured electrode, and the pick-up unit that can produce with test sample electrical effect when the electrochemical reaction is connected.Preferably, the material of this electrode system can be selected the conductive slurry-like material that is fit to screen painting, such as but not limited to carbon paste, gold size, elargol, carbon silver epoxy glue, volatility graphite or copper glue or its combination (printing again carbon paste after for example first printing elargol with wire mark), or the conductive slurry-like material of other any suitable screen painting.
Said electrical isolation separation layer herein, mean to have the formed thin layer of material of electrical insulation property preferably, the material of this electrical isolation separation layer can be selected electrical insulating property pulpous state material or the electrical insulating property adhesive tape that is fit to screen painting, the insulating tape of for example insullac of heated drying type or ultraviolet light drying type, or PVC or PET material.Preferably, to have thickness be 0.1 to 0.25 millimeter to this electric insulation layer.
According to of the present invention one better enforcement, this electrical isolation separation layer part is covered in on-chip electrode system, makes the electrode system zone that exposes not covering electric insulation layer, forms one and holds electrochemical electrode reaction zone and the vent port in sample solution space.
Said responding layer, mean to be positioned at above-mentioned electrochemical electrode reaction zone herein, contains the biological reagent with the measured matter reaction.Responding layer comprises enzyme and a kind of electron transfer mediator with the measured matter reaction, also contains simultaneously the composite thin layer of damping fluid, bonding agent and surfactant.Preferably, after the mode that this composite can apply or drip adds to this responding layer, dry under 60 ℃.
In the assay method and biology sensor of glucose in urine of the present invention, the above-mentioned working electrode of above-mentioned urine enrichment factor determination part and the voltage that applies between electrode is preferably greater than the voltage that equals water electrolysis, be preferably the scope of 1-5V, more preferably the scope of 1-3V.By applying the voltage more than or equal to water electrolysis voltage, can only depend on the concentrated electric current of urine with higher sensitivity determination, and can access the impact that is not subjected to existing other redox materials in urine, the stable electric current that does not rely on individual differences.In addition, application time is for example 1-10 second, 1-5 second more preferably.
For the purpose, technical scheme and the advantage that make the embodiment of the present invention is clearer, below in conjunction with embodiment and accompanying drawing, the embodiment of the present invention is described in further details.At this, illustrative examples of the present invention and explanation thereof are used for explanation the present invention, but not as a limitation of the invention.Can light facile modification on any person skilled in the art and change and all be contained in the appended scope of applying for a patent claim of this case.
Embodiment 1
The present embodiment provides the preparation example of an electric pole type Tes-Tape, thereby electrochemica biological sensor of the present invention and preparation method are described.As shown in Fig. 1-8:
At first, adopt screen printing technique to make counter electrode 2, working electrode 3, enrichment factor on the thick PET plastic substrate of 0.25mm and measure electrode 4, counter electrode 2, working electrode 3, enrichment factor are measured electrode 4 and are adopted the conductive carbon ink making of being bought by Acheson company.By serigraphy with electrically conductive ink after spreading finely dispersed printing ink thin layer on the PET plastic substrate, be put in 80 ℃ of baking ovens and dry 60min.
Secondly, adopt screen printing technique print insullac as electrical isolation separation layer 5 on the substrate of printed electrode system, insullac is by the purchase of Acheson company.By serigraphy with insullac after spreading finely dispersed printing ink thin layer on the PET plastic substrate, be put into ultraviolet dryer one minute oven dry.
Then, be used for the detection of urine glucose at electrode surface immobilized enzyme reaction film 7, responding layer only covers counter electrode, working electrode, does not measure electrode and do not cover enrichment factor.Each component and the percentage by weight thereof of the enzyme reaction solution of getting are as follows:
Get 4 μ L enzyme reaction solutions and be added drop-wise to electrode surface, 50 ℃ of oven dry 20min, standby.
Fixedly after enzyme membrane 7, with bonding partition 10, that surface encapsulation sheet 12, PET substrate 1 is bonding, bonding partition 10 is provided with siphon and leads sample groove 8, corresponding to the electrode reaction zone.
Use the urinary glucose determination of this biology sensor for example can followingly to implement.That is, at first collect urine with urine cup, on the other hand, above-mentioned biology sensor is arranged on supporting determinator, then make thief hatch 9 and the comes in contact with urine of the biology sensor that is arranged on determinator, by capillarity, urine is directed into biology sensor inside.Carry out in the steps below by the analysis that this biology sensor carries out.
(step 1: the detection of sample)
Apply voltage between two electrodes to electrode 2, working electrode 3, the current value that occurs by the importing along with urine changes to detect the importing of urine.In case confirmed the importing of urine, begun following step.The voltage that applies in step 1 for example is 0.1-0.5V.
(step 2: the mensuration of glucose)
After making glucose in urine and glucose oxidoreductase reaction certain hour, apply voltage between to electrode 2, working electrode 3, the amboceptor oxidation of the reducing condition that will produce on working electrode 3 by enzyme reaction detects its oxidation current.The reaction time of above-mentioned glucose and oxidoreducing enzyme is for example 1-30 second, 1-10 second more preferably.In step 2 apply voltage and application time is for example 0.1～1V, is preferably 0.2～0.5V, application time is for example 1-30 second, 1-10 second more preferably.
(step 3: the mensuration of enrichment factor)
Apply voltage between to electrode 2, urine enrichment factor detecting electrode 4, can detect the electric current that depends on enrichment factor thus, this enrichment factor is take the electrolytic oxidation reaction of urine composition as the basis.Being scaled enrichment factor by the electric current that detects can be by obtaining calibration curve or the calibration curve table carries out in advance.This correction can be used the enrichment factor of trying to achieve from the calibration curve of the electric current that makes in advance and enrichment factor, also can directly use the electric current that detects.Apply voltage and application time in step 3 are for example 1～5V, 2～3V more preferably, and application time is for example 1-30 second, 1-10 second more preferably.In this step, do not configure amboceptor on urine enrichment factor detecting electrode 4, and between working electrode 2 and enrichment factor detecting electrode 4, certain interval is arranged, do not configure the reagent such as amboceptor and only have urine in this gap, therefore the oxidation current that is not subjected to agents influence and depends on enrichment factor can be detected.This step 3 is preferably implemented after step 2 is completed.
(step 4: the correction of glucose in urine concentration)
By the enrichment factor that detects, proofread and correct the concentration of glucose that obtains in step 2 in step 3.This correction is preferably carried out as the basis take the calibration curve (containing correction card) that makes in advance.The concentration of glucose of proofreading and correct is shown by determinator or stores.As mentioned above, can not also to carry out the correction of glucose amount after formerly calculating enrichment factor, but directly use the electric current that depends on enrichment factor that detects in step 3 to come corrected glucose concentration.
Comparative example 1
This comparative example prepares biology sensor, except responding layer 7 had both covered electrode 2, working electrode 3, also covers enrichment factor and measures electrode 4, and other is identical with embodiment 1.
In the present embodiment, change in the scope of 1～5V applying voltage, measure the response current that enrichment factor is measured electrode.The correction of the mensuration of urine and glucose and glucose in urine concentration is identical with embodiment 1.The biology sensor that is used for this mensuration is made similarly to Example 1.Reagent layer 7 is configured on electrode 1, working electrode 2, is not disposed at enrichment factor and measures on electrode 3.The mensuration of response current value is also identical with embodiment 1.This measurement result is shown in each figure of Fig. 4-Fig. 8.Fig. 4-Fig. 8 represents with respect to the time dependent figure of response current value (nA) that applies voltage (V).
As shown in Figure 4, even 1V applies the response current that voltage also can detect the reflection enrichment factor, if but apply 2～5V, as Fig. 5～shown in Figure 8, can more clearly detect response current, most preferably as Fig. 5～apply as shown in Figure 6 situation of 2-3V.If apply 5V or more than, disorderly through waveform along with the time, but so long as in the short time after just applying, just can detect clearly the response current that reflects enrichment factor.In the present embodiment, change under certain condition applies voltage and detects take the electric current of enrichment factor as the basis, and can proofread and correct based on this glucose in urine concentration.
Claims (12)
1. the biology sensor that uses in the assay method of glucose in urine is characterized in that being divided into glucose in urine test section and enrichment factor test section; The glucose in urine test section has the 1st electrode system and reagent layer, and the mentioned reagent layer has oxidoreducing enzyme and the amboceptor of glucose in urine; The enrichment factor test section has working electrode and to electrode, does not configure amboceptor on above-mentioned working electrode, and configure amboceptor on to electrode; Described biology sensor comprises insulating substrate, namely has straight surface and the thin-layer tablet of electrical insulation characteristics; Comprise the electrode system that is positioned on insulating substrate, this electrode system is provided with three strip conductive electrodes, is respectively contrast electrode, working electrode, enrichment factor mensuration electrode; Comprise that the part is covered in the electric insulation layer on electrode system, make the electrode system zone that exposes not covering electric insulation layer, form one and hold electrochemical electrode reaction zone and the vent port in sample solution space; Comprise the responding layer that is positioned at the electrochemical electrode reaction zone, this responding layer comprises at least a oxidoreducing enzyme, a kind of electron mediator, also comprise damping fluid, bonding agent and surfactant, responding layer only covers contrast electrode, working electrode, does not measure electrode and do not cover enrichment factor.
2. the biology sensor that uses in the assay method of glucose in urine as claimed in claim 1 is characterized in that described insulating substrate is selected from the following group who forms: polyvinyl chloride (PVC) sheets, glass fiber sheets, polyester, bakelite sheet, PET sheet, PC sheet, PP sheet, PE sheet, PA sheet, PS sheet, glass sheet and potsherd.
3. the biology sensor that uses in the assay method of glucose in urine as claimed in claim 1, it is characterized in that described electrode system is to cover so that electric insulation layer is local, make and be exposed to electrical isolation separation layer one end and form respectively working electrode, reference electrode, enrichment factor and measure electrode and be connected with the electrochemical electrode reaction zone, the other end forms the joint that working electrode, reference electrode, enrichment factor are measured electrode, and the pick-up unit that produces electrical effect with test sample when the electrochemical reaction is connected.
4. the biology sensor that uses in the assay method of glucose in urine as claimed in claim 3, it is characterized in that the material selection of described electrode system is fit to the conductive slurry-like material of screen painting, for example carbon paste, gold size, elargol, carbon silver epoxy glue, volatility graphite or copper glue or its combination, or the conductive slurry-like material of other suitable screen painting.
5. the biology sensor that uses in the assay method of glucose in urine as claimed in claim 1, it is characterized in that the material selection of described electric insulation layer is fit to electrical insulating property pulpous state material or the electrical insulating property adhesive tape of screen painting, as the insullac of heated drying type or ultraviolet light drying type, or the insulating tape of PVC or PET material.
6. the biology sensor that uses in the assay method of glucose in urine as claimed in claim 5, the thickness that it is characterized in that described electric insulation layer are 0.1 to 0.25 millimeter.
7. the biology sensor that uses in the assay method of glucose in urine as claimed in claim 1, it is characterized in that described responding layer comprises enzyme and a kind of electron transfer mediator that reacts with measured matter, also contains the composite thin layer of damping fluid, bonding agent and surfactant simultaneously.
8. the biology sensor that uses in the assay method of glucose in urine as claimed in claim 7, the composite thin layer that it is characterized in that described responding layer are after the mode that applies or drip adds to this responding layer, and be dry under 60 ℃.
9. the assay method of glucose in urine, the steps include:
A under the existence of amboceptor, utilizes oxidoreducing enzyme that the glucose oxidase in urine is reduced;
B detects by electrode system oxidation current or the reduction current that produce this moment, take above-mentioned current value as the basic calculation concentration of glucose;
C measures the conductivity in above-mentioned urine and utilizes this conductivity to proofread and correct the correcting process of urine glucose concentration; The assay method of conductivity is: prepare to contain working electrode and to the electrode system of electrode, do not configure amboceptor on the working electrode in above-mentioned two electrodes, and configure amboceptor on to electrode; Urine is directed into above-mentioned electrode system, under this state, above-mentioned electrode system is applied voltage, detect thus and flow to above-mentioned two interelectrode oxidation current or reduction currents, take this current value as the basic calculation conductivity value.
10. the assay method of the glucose in urine of glucose in urine as claimed in claim 9, is characterized in that described electron mediator is the potassium ferricyanide, and its content is that 3% (m/m) is to 20% (m/m).
11. the assay method of the glucose in urine of glucose in urine as claimed in claim 9 is characterized in that described damping fluid is a kind of in the group that forms of phosphate buffer, MES damping fluid, citrate buffer solution; Described bonding agent adopts one or more in starch, dextrin, gelatin, hydroxyethyl cellulose, carboxymethyl cellulose, sodium alginate; Described surfactant is TritonX-100 or TW-20.
12. the assay method of the glucose in urine of glucose in urine as claimed in claim 9 is characterized in that described oxidoreducing enzyme is glucose oxidase or glucose dehydrogenase.
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Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
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| CN104535753A (en) * | 2014-12-05 | 2015-04-22 | 方金 | Micro urine sugar detection test paper and detection method thereof |
| CN104949966A (en) * | 2014-03-26 | 2015-09-30 | 爱科来株式会社 | Reducing power analysis method and reducing power analysis reagent |
| CN105722453A (en) * | 2012-08-06 | 2016-06-29 | 威里利生命科学有限责任公司 | Contact lenses having two-electrode electrochemical sensors |
| CN106770542A (en) * | 2016-11-25 | 2017-05-31 | 深圳大学 | A kind of noninvasive dynamics monitoring test paper and preparation method thereof |
| CN108760856A (en) * | 2018-05-30 | 2018-11-06 | 杭州点壹下通讯科技有限公司 | A kind of urine detection method based on intelligent closestool |
| CN109085218A (en) * | 2018-01-19 | 2018-12-25 | 上海荒岛科技有限公司 | A kind of reagent and electrochemical sensor |
| CN109239158A (en) * | 2018-10-12 | 2019-01-18 | 京东方科技集团股份有限公司 | Glucose in urine test block and glucose in urine test method |
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Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN105722453A (en) * | 2012-08-06 | 2016-06-29 | 威里利生命科学有限责任公司 | Contact lenses having two-electrode electrochemical sensors |
| CN104949966A (en) * | 2014-03-26 | 2015-09-30 | 爱科来株式会社 | Reducing power analysis method and reducing power analysis reagent |
| CN104949966B (en) * | 2014-03-26 | 2018-01-02 | 爱科来株式会社 | The analysis method of reducing power and the analytical reagent of reducing power |
| CN104330448A (en) * | 2014-10-31 | 2015-02-04 | 桂林中辉科技发展有限公司 | High-sensitivity electrode type uric acid test paper and manufacturing method thereof |
| CN104535753A (en) * | 2014-12-05 | 2015-04-22 | 方金 | Micro urine sugar detection test paper and detection method thereof |
| CN106770542A (en) * | 2016-11-25 | 2017-05-31 | 深圳大学 | A kind of noninvasive dynamics monitoring test paper and preparation method thereof |
| CN109085218A (en) * | 2018-01-19 | 2018-12-25 | 上海荒岛科技有限公司 | A kind of reagent and electrochemical sensor |
| WO2019214496A1 (en) * | 2018-05-05 | 2019-11-14 | 深圳市贝沃德克生物技术研究院有限公司 | Biosensor and method for detecting glucose concentration in diabetes biomarker |
| CN108760856A (en) * | 2018-05-30 | 2018-11-06 | 杭州点壹下通讯科技有限公司 | A kind of urine detection method based on intelligent closestool |
| CN109239158A (en) * | 2018-10-12 | 2019-01-18 | 京东方科技集团股份有限公司 | Glucose in urine test block and glucose in urine test method |
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