CN102419366B - Dry chemical quantitative test strip with interference removal, alanine transaminase or aspartate transaminase quantitative test strip and test method - Google Patents
Dry chemical quantitative test strip with interference removal, alanine transaminase or aspartate transaminase quantitative test strip and test method Download PDFInfo
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- CN102419366B CN102419366B CN201110240460.8A CN201110240460A CN102419366B CN 102419366 B CN102419366 B CN 102419366B CN 201110240460 A CN201110240460 A CN 201110240460A CN 102419366 B CN102419366 B CN 102419366B
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Abstract
The invention discloses a dry chemical quantitative test strip with interference removal, comprising a substrate, a sampling region which is provided with a fixed interference removing reagent and arranged on the substrate, and a promoter region which is provided with a fixed test reaction promoter and a fixed signal supplying reagent and is arranged on the substrate, wherein, the sampling region and the promoter region are distributed on the test strip under the conditions that: the sampling region keeps non-contact with the promoter region in normal state, and the sampling region contacts with the promoter region in test state. The test method comprises the following steps: adding a sample liquid to the tested in the sampling region, redissolving the substance fixed on the sampling region in the sample liquid to the tested, carrying out reaction for removing interference; then contacting the promoter region with the sampling region, redissolving the substance fixed on the promoter region, and carrying out reaction for test; carrying out signal reaction on the reaction product, and according to the tested signal value, calculating the content of biological enzyme to be tested. The test method has the advantages of simple operation, low test cost, accurate test result, and high test precision, and can be concretely applied for the quantitative test of alanine transaminase or aspartate transaminase.
Description
Technical field
The present invention relates to a kind of dry chemical test examination bar and method of testing, relate in particular to a kind of dry chemical test examination bar and detection method of transaminase.
Background technology
Chaff interference in dry chemical test is a lot, and such as the interference of the uric acid in redox reaction mensuration, ascorbic acid etc., the pyruvic acid during transaminase detects disturbs, and especially with the pyruvic acid in transaminase, disturbs comparatively serious.
The kind of transaminase is a lot, wherein important with glutamic-pyruvic transaminase (GPT/ALT) and glutamic-oxalacetic transaminease (GOT/AST).Glutamic-pyruvic transaminase claims again alanine aminotransferase, is the transamination between catalysis glutamic acid and pyruvic acid; Glutamic-oxalacetic transaminease claims again AST, is the transamination between catalysis glutamic acid and oxaloacetic acid.Clinically, the mensuration of these two kinds of transaminases in whole blood, serum, blood plasma, tissue fluid, is very important index in heart disease, muscle disease, especially diagnosing hepatism.
Measure clinically the method for above-mentioned two kinds of transaminases, it is mainly the method for using the liquid reagent of Biochemical Analyzer, the method is to utilize lactic dehydrogenase, malic dehydrogenase catalysis glutamic-pyruvic transaminase product pyruvic acid and the dehydrogenation of glutamic-oxalacetic transaminease product oxaloacetic acid respectively, consume nicotinamide adenine dinucleotide (NADH), cause that light absorption value is in the variation of 340nm.This rate of change and glutamic-pyruvic transaminase, the active proportional relation of glutamic-oxalacetic transaminease, thereby the enzymatic activity of drawing.
Reaction mentioned above is as follows:
Glutamic-pyruvic transaminase:
Glutamic-oxalacetic transaminease:
The method is consuming time, complicated operation, need to be than relatively large instrument; And in the method, along with the carrying out of reaction, NADH constantly consumes, and the absorbance of 340nm constantly reduces.Because the reason of the aspects such as instrument, initial NADH can not be too high, and this is affected the test specification of the method, easily occurs false negative when the high value sample of test transaminase.
Dry chemical method has conveniently, flexibly, pollute the features such as little, easy and simple to handle.At present, dry chemical method roughly can be divided into following a few class: take the electrochemical method (as US Patent No. 6565738) that Abbott Laboratories are representative.The δ-bilirubin that Fuji, Kodak, Johnson & Johnson are representative (as US Patent No. 4897347, US5508173, US5462858).The cross flow dry chemical method that the Roche of take is representative (as US Patent No. 4591553, US5508173, US4665023).Yet, all there is part defect in preceding method, such as Abbott Laboratories' electrochemical process test glutamic-pyruvic transaminase, adopts glucose oxidation enzyme process oxidation reaction (1) to produce glutamic acid, but exist endogenic alanine to disturb (more more common than endogenic pyruvic acid), affect test result.And the dry chemical method that the δ-bilirubin that Fuji is representative, Roche are representative all adopts coupling Pyruvate oxidase method.Adopt the course of reaction of coupling pyruvate oxidation enzyme method as follows:
Glutamic-pyruvic transaminase, coupling after reaction (1)
Glutamic-oxalacetic transaminease, coupling after reaction (3)
According to patent US4665023, as shown in Figure 1, blood sample is added on diffusion layer 1 structure of its examination bar, and after hemofiltration film 3 hemofiltrations, blood plasma flows to and flows on pad 5.1min after application of sample, presses down transparent protective seam 9, and reagent layer 6 and enzyme pad 4 contact with the pad that flows.The pyruvate oxidase, the peroxidase that are fixed on developer, the zymolyte on reagent layer 6 and are fixed on enzyme pad 4 redissolve in sample, thereby produce detectable color signal.
From test philosophy, can clearly find out, there is the interference of endogenous pyruvic acid in the method.The pyruvic acid of color signal is provided, and existing enzymatic produces, and also has the endogenous pyruvic acid itself existing in sample.And employing rate method (i.e. the rate of change of test colour developing), remove endogenous and disturb the optical instrument that needs more complicated, usually adopt the optical device such as integrating sphere, and the normal value of transaminase is generally very low, glutamic-pyruvic transaminase is 5U/L～40U/L, glutamic-oxalacetic transaminease is 8U/L～40U/L, and the normal reference value of pyruvic acid is relatively high, it is 65 μ mol/L (being equivalent to the pyruvic acid that 65U/L transaminase produces at 1min), within the shorter dry chemical test duration (2min～3min), the rate of change of chromogenic reagent also can be subject to the impact of endogenous pyruvic acid to a certain extent, easily there is false positive results.
Summary of the invention
The technical problem to be solved in the present invention is to overcome the deficiencies in the prior art, provide a kind of measurement result accurately, preparation is simple, testing cost is low, detect dry chemical quantitative test examination bar that easy to operate removal disturbs and especially for the quantitative test examination bar of paddy third or glutamic-oxalacetic transaminease, also provides the quantitative test examination bar of a kind of easy and simple to handle, testing cost is lower, testing result is accurate, accuracy of detection is high this paddy third of use or glutamic-oxalacetic transaminease quantitatively to detect the method for blood two-story valley third or glutamic-oxalacetic transaminease.
For solving the problems of the technologies described above, the technical scheme that the present invention proposes is a kind of dry chemical quantitative test examination bar of removing interference, described test examination bar comprises end liner, on described end liner, arrange one and be fixed with the sample application zone of disturbing reagent, on described end liner, also arrange one and be fixed with the promoter region that signal provides reagent and test reaction to start reagent, described sample application zone and promoter region are that the mode that keeps in the normal state noncontact, can contact with each other under test mode is laid on described test examination bar.Described going disturbs reagent mainly to refer to the reaction reagent that can remove endogenous interfering material in testing sample, described signal provides reagent to refer to reaction that can be based on endogenous interfering material and then the corresponding reagent of background signal is provided, described test reaction start reagent comprise preferably can the reaction of startup analysis thing reaction reagent, signal produce some biology enzyme reagent that reagent and analytical reactions need etc.
The first as the dry chemical quantitative test examination bar that above-mentioned removal is disturbed is improved, described sample application zone is directly fixed on described end liner, described promoter region is connected on described end liner by a bonding piece, and this bonding piece has most of region that enough height make described promoter region and is suspended in described sample application zone top.In this preferred technical scheme, because promoter region is suspended in sample application zone top, therefore under normal conditions, sample application zone and promoter region are to exist with non-contacting state; When detecting with this test examination bar, only promoter region need be pressed downwards, just can make promoter region and sample application zone keep in touch, to start corresponding detection reaction.
The second as the dry chemical quantitative test examination bar that above-mentioned removal is disturbed improves, described sample application zone and promoter region are separated by a separate layer, this separate layer is set to the connected mode of drawable formula, so that this separate layer pulls out described sample application zone and promoter region after test examination bar, can keep in touch.In this preferred technical scheme, because promoter region and sample application zone are divided on the pros and cons of separate layer, therefore under normal conditions, between sample application zone and promoter region, by separate layer, isolate and exist with non-contacting state; When detecting with this test examination bar, only need this separate layer to pull out separately test examination bar, just can make promoter region and sample application zone keep in touch, to start corresponding detection reaction.
The third improvement as the dry chemical quantitative test examination bar that above-mentioned removal is disturbed, described sample application zone can directly be fixed on described end liner, described promoter region is articulated with one end of described end liner by an articulated elements, the rotating shaft of described articulated elements is axially parallel with surface level or vertical, so that promoter region can be kept in touch with described sample application zone after rotating.When rotating shaft is axially parallel with surface level, described promoter region be by the upset realization in perpendicular and sample application zone by noncontact to contacting; When rotating shaft is axially vertical with surface level, described promoter region be by the rotation realization in surface level and sample application zone by noncontact to contacting, now the height of articulated elements should be suitable, so that promoter region can just touch sample application zone after rotating translation.
The 4th kind of improvement as the dry chemical quantitative test examination bar that above-mentioned removal is disturbed, described sample application zone and promoter region are separately fixed at the zones of different (mutually keeping certain distance) of described end liner the same face, end liner in the middle of described sample application zone and promoter region is provided with a crease line, so that described end liner can make promoter region and sample application zone keep in touch after crease line bending.
In the dry chemical quantitative test examination bar that above-mentioned various removals are disturbed; promoter region can arrange a reagent layer; its effect is to start transamination reaction; preferred material is polycarbonate or nylon; also material on reagent layer directly can be fixed in fibrous material as reagent layer; now can on reagent layer, set up as required the protective seam of layer of transparent, described protective seam is preferably made of transparent polycarbonate or pvc material.
In the dry chemical quantitative test examination bar that above-mentioned various removals are disturbed, structure specific to sample application zone and promoter region, described sample application zone can be according to the needs of detected sample, whole or several in sample flow pad, diffusion layer, sample pad, hemofiltration film, enzyme pad form, described sample application zone preferably includes and is fixed on the mobile pad on end liner and is fixed on the enzyme pad flowing on pad, and described enzyme pad is provided with a kind of or stack in hemofiltration film, sample pad, diffusion layer wherein multiple is set.For example, on described enzyme pad, can superpose successively hemofiltration film, sample pad and diffusion layer are set, and for example, when detecting serum with this test examination bar, just can cancel hemofiltration rete.Described promoter region also can be according to the needs of detected sample, comprise one or more in reagent layer, enzyme pad, protective seam.
In the dry chemical quantitative test examination bar that above-mentioned removal is disturbed, described end liner can adopt (transparent or opaque) macromolecule polymeric material (as tygon, Polyvinylchloride, polystyrene, polyester etc.) to make, but preferably adopt Polyvinylchloride (PVC), polycarbonate, polyamide macromolecule polymeric material to make, the thickness of end liner is preferably 50 μ m～300 μ m.The material that described mobile pad can adopt comprises glass fibre, dacron, nitrocellulose filter, polysulfone membrane, poly (ether sulfone) film etc., preferably adopt glass fibre, nitrocellulose filter, dacron film or filter paper to make, its thickness is preferably 0.04mm～0.2mm.Described enzyme pad preferably adopts glass fibre, cellulose filter paper, dacron or filter paper to make.The effect of described hemofiltration film is to filter haemocyte, if sample to be tested is blood plasma, serum, this layer can be set, and its material can be the special-purpose hemofiltration film that various glass fibre and some producers provide, and preferably adopts asymmetric polysulfone membrane or glass fibre to make.Described sample pad act as maintenance sample, do not allow sample overflow (inessential setting), its material can be glass fibre, dacron etc., preferably adopts glass fibre to make.The effect of described diffusion layer is to make testing sample diafiltration (inessential setting) in lower floor equably, the screen cloth form that preferably adopts dacron or nylon fiber to make, screen cloth aperture is preferably 40 order～150 orders, fibre diameter is preferably 100 μ m～500 μ m, and screen cloth is preferably hydrophilic or hydrophilic treatment.
As a total technical conceive, the present invention also provides the method for a kind of use dry chemical quantitative test that above-mentioned removal is disturbed examination bar detection of biological enzyme, comprise the following steps: the sample application zone of first analyte sample fluid being added to described test examination bar, in described sample application zone, fixing going disturbs reagent to redissolve in analyte sample fluid, the endogenous interfering material containing in described analyte sample fluid disturbs reagent to start to react with going in analyte sample fluid, after one section of hatching process, (conventionally keep 1min), make again the promoter region of described test examination bar contact with sample application zone, on promoter region, fixing signal provides reagent and test reaction startup reagent to redissolve in analyte sample fluid, and start the test reaction of startup analysis thing, the product generating after test reaction starts provides reagent to carry out signal reaction with described signal again, the signal value recording according to signal reaction, calculate the content of biology enzyme to be measured.In above-mentioned detection method, the discernible signal that described signal reaction produces comprises and a kind of in color signal, fluorescence signal, chemiluminescence signal most preferably is color signal, and especially adopting absorbing wavelength is the color signal of 600nm～700nm.
As a total technical conceive, the present invention also provides a kind of paddy third or glutamic-oxalacetic transaminease quantitative test examination bar, this quantitative test examination bar is to be made by above-mentioned dry chemical quantitative test examination bar, test reaction startup reagent fixing on described promoter region mainly refers to the substrate that can start transamination reaction, in described sample application zone, fixing going disturbs reagent to comprise pyruvate oxidase and hydrogen peroxidase (CAT), in described promoter region or sample application zone, be also fixed with peroxidase (POD), on described promoter region, fixing signal provides reagent to comprise to be applicable to the chromogenic substrate of described peroxidase, fluorogenic substrate or chemical luminous substrate.In addition situation per sample, can add the materials such as ascorbic acid oxidase or urate oxidase in described sample application zone.
As a total technical conceive, the method that the present invention also provides the above-mentioned paddy of a kind of use third or glutamic-oxalacetic transaminease quantitative test examination bar to detect paddy third or glutamic-oxalacetic transaminease (does not arrange hemofiltration film, the situation of sample pad and diffusion layer), comprise the following steps: the sample application zone of first analyte sample fluid being added to described test examination bar, fixing pyruvate oxidase in described sample application zone, hydrogen peroxidase all redissolves in analyte sample fluid, the endogenous interfering material pyruvic acid containing in described analyte sample fluid and pyruvate oxidase and the hydrogen peroxidase in analyte sample fluid start disturbance reponse, after one section of hatching process, make again the promoter region of described test examination bar contact with sample application zone, on promoter region, fixing substrate redissolves in analyte sample fluid, peroxidase fixing in promoter region or sample application zone has also redissolved in analyte sample fluid, now start to start transamination reaction, the pyruvic acid generating after transamination reaction starts participates in coupling reaction, the hydrogen peroxide generating after coupling reaction and described chromogenic substrate carry out chromogenic reaction, according to the reflectance value of chromogenic substrate colour developing and determine the content of paddy third or glutamic-oxalacetic transaminease by end-point method or rate method.
As a total technical conceive, the present invention also provides another kind of paddy third or glutamic-oxalacetic transaminease quantitative test examination bar, described quantitative test examination bar is had specific sample application zone structure and (in this sample application zone, is comprised enzyme pad by above-mentioned, hemofiltration film, sample pad and diffusion layer) dry chemical quantitative test examination bar makes, test reaction startup reagent fixing on described promoter region mainly refers to the substrate that can start transamination reaction, in described sample application zone, fixing going disturbs reagent to comprise pyruvate oxidase and hydrogen peroxidase, in described promoter region or sample application zone, be also fixed with peroxidase, on described promoter region, fixing signal provides reagent to comprise to be applicable to the chromogenic substrate of described peroxidase, fluorogenic substrate or chemical luminous substrate, described enzyme pad is provided with hemofiltration film, and described hemofiltration film is provided with sample pad, and described sample pad is provided with diffusion layer.In diffusion layer, sample pad and hemofiltration film, can add buffer system, surfactant, anti-haemolysis reagent (as carbohydrate), salt (as sodium chloride) etc.
As a total technical conceive, the present invention also provides the method that detects paddy third or glutamic-oxalacetic transaminease with the paddy third of above-mentioned the second or glutamic-oxalacetic transaminease quantitative test examination bar, comprise the following steps: first analyte sample fluid is added drop-wise on the diffusion layer of described test examination bar sample application zone, make analyte sample fluid flow through successively sample pad and hemofiltration film infiltrate gradually on described enzyme pad, pyruvate oxidase and hydrogen peroxidase fixing in described sample application zone all redissolve in analyte sample fluid, the endogenous interfering material pyruvic acid containing in described analyte sample fluid and pyruvate oxidase and the hydrogen peroxidase in analyte sample fluid start disturbance reponse, after one section of hatching process, make again the promoter region of described test examination bar contact with sample application zone, on promoter region, fixing substrate redissolves in analyte sample fluid, peroxidase fixing in promoter region or sample application zone has also redissolved in analyte sample fluid, now start to start transamination reaction, the pyruvic acid generating after transamination reaction starts participates in coupling reaction, the hydrogen peroxide generating after coupling reaction and described chromogenic substrate carry out chromogenic reaction, according to the reflectance value of chromogenic substrate colour developing and determine the content of paddy third or glutamic-oxalacetic transaminease by end-point method or rate method.
In above-mentioned paddy third or glutamic-oxalacetic transaminease quantitative test examination bar, the concentration of described pyruvate oxidase is preferably 30KU/L～300KU/L.In above-mentioned paddy third or glutamic-oxalacetic transaminease quantitative test examination bar, the concentration of described peroxidase is preferably 30KU/L～200KU/L.In above-mentioned paddy third or glutamic-oxalacetic transaminease quantitative test examination bar, described catalatic concentration is 1KU/L～200KU/L.In above-mentioned paddy third or glutamic-oxalacetic transaminease quantitative test examination bar, the concentration of described chromogenic substrate, fluorogenic substrate or chemical luminous substrate is preferably 1mM～50mM.In technique scheme, selecting of developer can be determined (having the american documentation literatures such as US4591553, US5274095, US5162200, US4666023 mentions) according to actual needs.If developer system only includes a kind of component, this component is fixed on the reagent layer of promoter region, if developer system comprises various ingredients, has at least a kind of component to be fixed on the reagent layer of promoter region.
In above-mentioned paddy third or glutamic-oxalacetic transaminease quantitative test examination bar, the substrate of described startup transamination reaction preferably includes α-ketoglutaric acid, and the concentration of described α-ketoglutaric acid is preferably 5mM～300mM; The substrate of described startup transamination reaction also preferably includes for detection of the ALANINE of glutamic-pyruvic transaminase or for detection of the L-ASPARTIC ACID of glutamic-oxalacetic transaminease.In fact, if be fixed on the substrate of the startup transamination reaction of promoter region, comprised α-ketoglutaric acid, materials such as ALANINE, L-ASPARTIC ACID also can be fixed in sample application zone so.
As nothing is mentioned especially, on pilot scale bar of the present invention, the concentration of accompanying material all refers to that routine chromogenic agent described above refers to when preparation examination bar for soaking the concentration of this selected chromogenic reagent solution of carrier film in preparation concentration for this selected substance solution of wetting carrier film during this examination bar.
In sum, in test examination bar of the present invention, reaction system can comprise buffer system, transamination reaction substrate, anti-haemolysis reagent, pyruvate oxidase (concentration is preferably 30KU/L～300KU/L), peroxidase (concentration is preferably 30KU/L～200KU/L), transaminase activator, diphosphothiamine (TPP, concentration is preferably 0.4mg/ml～1mg/m1), flavin adenine dinucleotide (FAD) (FAD, concentration is preferably 0.1mg/ml～0.2mg/m1), surfactant, developer, pyruvic acid activator be (as Mg
2+, Mn
2+, be preferably the Mg of 0.01M～0.1M
2+), paddy third, glutamic-oxalacetic transaminease activator (being preferably the P5P of 1mM～500mM), stabilizing agent etc.Wherein, the activity that comprises selection, concentration and the enzyme of each non-key component of buffer system all can be determined (can with reference to american documentation literatures such as US4271265, US4666832, US4591553) according to prior art voluntarily by those skilled in the art.The materials such as TPP, FAD, surfactant, activator all can be fixed in sample application zone.
Above-mentioned paddy third of the present invention or glutamic-oxalacetic transaminease quantitative test examination bar and detection method are mainly based on following principle: on the basis of coupling pyruvate oxidase, by adding again hydrogen peroxidase can very effectively remove the interference of endogenous material pyruvic acid, because the H that endogenous interfering material pyruvic acid produces
2o
2(referring to above-mentioned reaction (5)) are all got rid of by following reaction (8) by hydrogen peroxidase, therefore participate in the H of follow-up chromogenic reaction
2o
2(referring to above-mentioned reaction (6)) are all that the product by transamination reaction is transformed.
Compared with prior art, the invention has the advantages that: by adopt test of the present invention examination bar and detection method can be more accurately, easily glutamic-pyruvic transaminase or glutamic-oxalacetic transaminease are carried out to qualitative detection and quantitative test, and test examination bar of the present invention is simple in structure, be convenient to make, detection method is easy to operate, quick, without newly-increased other instruments or equipment, meet the requirement of the diagnosis (POCT) of bedside, be applicable to the uses such as hospital emergency, ward, user oneself, community hospital, have broad application prospects.
Accompanying drawing explanation
Fig. 1 is the structural representation of dry chemical quantitative test examination bar in prior art.
The dry chemical quantitative test that goes interference that Fig. 2 provides for the embodiment of the present invention 1 tries the structural representation of bar.
The dry chemical quantitative test that goes interference that Fig. 3 provides for the embodiment of the present invention 2 tries the structural representation of bar.
The dry chemical quantitative test that goes interference that Fig. 4 provides for the embodiment of the present invention 3 tries the structural representation of bar.
The dry chemical quantitative test that goes interference that Fig. 5 provides for the embodiment of the present invention 4 tries the structural representation of bar.
The dry chemical quantitative test that goes interference that Fig. 6 provides for the embodiment of the present invention 5 tries the structural representation of bar.
The dry chemical quantitative test that goes interference that Fig. 7 provides for the embodiment of the present invention 6 tries the structural representation (overlooking) of bar.
The dry chemical quantitative test that goes interference that Fig. 8 provides for the embodiment of the present invention 6 tries the structural representation (main looking) of bar.
The dry chemical quantitative test that goes interference that Fig. 9 provides for the embodiment of the present invention 2 tries another variation of bar.
Figure 10 is the linear regression equation of 2 pairs of glutamic-pyruvic transaminase of comparative example of providing in the embodiment of the present invention 7 foundation while carrying out quantitative test.
Figure 11 is the comparison diagram of reflectance value in the embodiment of the present invention 7 and comparative example 1.
Figure 12 is the linear regression equation of foundation while glutamic-oxalacetic transaminease being carried out to quantitative test in comparative example 4 of the present invention.
Figure 13 is the comparison diagram of reflectance value in the embodiment of the present invention 8 and comparative example 3.
Marginal data:
1, diffusion layer; 2, sample pad; 3, hemofiltration film; 4, enzyme pad; 5, flow and pad; 6, reagent layer; 7, end liner; 8, bonding piece; 9, protective seam; 10, articulated elements; 11, crease line; 12, separate layer; 13, sample application zone; 14, promoter region.
Embodiment
Below in conjunction with Figure of description and specific embodiment, the invention will be further described.
Embodiment 1:
The dry chemical quantitative test examination bar that a kind of removal is as shown in Figure 2 disturbed, this test examination bar comprises end liner 7, on end liner 7, arrange one and be fixed with the sample application zone 13 (specifically comprising diffusion layer 1, sample pad 2, hemofiltration film 3, enzyme pad 4 and the pad 5 that flows in the present embodiment) of disturbing reagent, sample application zone 13 is directly fixed on end liner 7; On end liner 7, also arranging one is fixed with test reaction and starts the promoter region 14 that reagent and signal provide reagent, promoter region 14 comprises a reagent layer 6, reagent layer 6 is connected to one end of end liner 7 by a bonding piece 8, this bonding piece 8 has the top that most of region that enough height make reagent layer 6 is suspended in sample application zone 13.Sample application zone 13 and promoter region 14 keep noncontact in the normal state, under test mode, can itself and sample application zone 13 (being specially the mobile pad 5 of sample application zone 13) be contacted with each other by downward started by press district 14 (reagent layer 6).
In the present embodiment, sample application zone 13 comprises the mobile pad 5 being fixed on end liner 7, flow and pad one end immobilized enzyme pad 4 of 5, the top of enzyme pad 4 sets gradually hemofiltration film 3, sample pad 2 and diffusion layer 1, and a side of diffusion layer 1, sample pad 2, hemofiltration film 3, enzyme pad 4 and the pad 5 that flows interconnects by bonding piece 8, and is finally fixed on end liner 7.The reagent layer 6 of promoter region 14 is specifically arranged on mobile pad 5 tops of sample application zone 13.Go to disturb reagent to be fixed on enzyme pad 4 or flow on pad 5, test reaction starts reagent and signal provides reagent to be fixed on reagent layer 6.
The method of the dry chemical quantitative test examination bar detection of biological enzyme that the above-mentioned removal of a kind of use is disturbed (is applicable to whole blood test, serum, blood plasma etc.), comprise the following steps: first analyte sample fluid (as: whole blood) is added on the diffusion layer 1 of test examination bar sample application zone 13, through sample pad 2, hemofiltration film 3 is to filter haemocyte, blood plasma redissolves enzyme pad 4 or flow and remove to disturb reagent on pad 5, finally be penetrated into and flow on pad 5, the going of the endogenous interfering material containing in analyte sample fluid and redissolution disturbs reagent to start disturbance reponse, after the hatching process of 1min left and right, make again the reagent layer 6 of promoter region 14 and the mobile pad 5 of sample application zone 13 of test examination bar contact, test reaction startup reagent (comprising reaction substrate) and signal fixing on promoter region 14 provide the redissolution such as reagent in analyte sample fluid, and start the test reaction of startup analysis thing, the product generating after test reaction starts provides reagent to carry out signal reaction with signal again, the reflectance value recording according to signal reaction, calculate the content of biology enzyme to be measured.
Embodiment 2:
The dry chemical quantitative test examination bar that a kind of removal is as shown in Figure 3 disturbed, this test examination bar comprises end liner 7, on end liner 7, arrange one and be fixed with the sample application zone 13 (specifically comprising diffusion layer 1, sample pad 2, hemofiltration film 3, enzyme pad 4 and the pad 5 that flows in the present embodiment) of disturbing reagent, sample application zone 13 is directly fixed on end liner 7; On end liner 7, also arranging one is fixed with test reaction and starts the promoter region 14 that reagent and signal provide reagent, promoter region 14 comprises a reagent layer 6, reagent layer 6 is connected to one end of end liner 7 by a bonding piece 8, this bonding piece 8 has the top that most of region that enough height make reagent layer 6 is suspended in sample application zone 13.Sample application zone 13 and promoter region 14 keep noncontact in the normal state, under test mode, can itself and sample application zone 13 (specifically referring to the enzyme pad 4 in sample application zone 13) be contacted with each other by downward started by press district 14 (reagent layer 6).
In the present embodiment, sample application zone 13 comprises the mobile pad 5 being fixed on end liner 7, one end of mobile pad 5 (near one end of promoter region 14) is fixed with enzyme pad 4, the other end is fixed with hemofiltration film 3, the top of hemofiltration film 3 is disposed with sample pad 2 and diffusion layer 1, and a side of diffusion layer 1, sample pad 2, hemofiltration film 3 and the pad 5 that flows interconnects by bonding piece 8, and is finally fixed on end liner 7.The reagent layer 6 of promoter region 14 is specifically arranged on enzyme pad 4 tops of sample application zone 13.Go to disturb reagent to be fixed on enzyme pad 4 or flow on pad 5, test reaction starts reagent and signal provides reagent to be fixed on reagent layer 6.
Therefore, than embodiment 1, embodiment 2 makes an adjustment the position of enzyme pad 4 and optimize, be about to enzyme pad 4 and be fixed on pad 5 one end near promoter region 14 of flowing, when detecting, by downward started by press district 14, can make it first contact with enzyme pad 4, the deviation that the difference that the present embodiment can avoid enzyme to discharge well causes.In addition, enzyme pad 4 also can be arranged on the below of reagent layer 6, as shown in Figure 9, becomes a part for promoter region 14, now goes to disturb reagent to be fixed on and flows on pad 5.
Embodiment 3:
The dry chemical quantitative test examination bar that removal is as shown in Figure 4 disturbed, the test providing than embodiment 1 examination bar, the test examination bar of the present embodiment is not established enzyme pad 4, and remaining structure and application mode are identical with embodiment 1.Owing to not establishing enzyme pad 4, the material (for example various catalytic reaction enzymes) being originally fixed on enzyme pad 4 for example can directly be fixed on, in other structures (pad 5 flows) of sample application zone 13.
Embodiment 4:
The dry chemical quantitative test examination bar that a kind of removal is as shown in Figure 5 disturbed, this test examination bar comprises end liner 7, on end liner 7, arrange one and be fixed with the sample application zone 13 (comprising flow pad 5, enzyme pad 4, hemofiltration film 3, sample pad 2 and diffusion layer 1 etc.) of disturbing reagent, sample application zone 13 is directly fixed on end liner 7; On end liner 7, also arrange one and be fixed with test reaction and start the promoter region 14 that reagent and signal provide reagent, promoter region 14 comprises a reagent layer 6, and reagent layer 6 is also directly fixed on end liner 7, and sample application zone 13 and promoter region 14 isolate mutually by a separate layer 12.Separate layer 12 is set to the connected mode of drawable formula, and sample application zone 13 and promoter region 14 keep noncontact in the normal state, under test mode, can the sample application zone 13 that be positioned at its positive and negative be kept in touch mutually with promoter region 14 by detaching separate layer 12.
In the present embodiment, mobile pad 5 parts for sample application zone 13 are directly fixed on end liner 7, a part is positioned at separate layer 12 tops, and the top, one end of the pad 5 that flows is provided with enzyme pad 4, and the top of enzyme pad 4 is disposed with hemofiltration film 3, sample pad 2 and diffusion layer 1.Go to disturb reagent to be fixed on enzyme pad 4, test reaction starts reagent and signal provides reagent to be fixed on reagent layer 6.
The test examination bar of the present embodiment for detection of time method of operating similar to embodiment 1, first analyte sample fluid (whole blood) is added on the diffusion layer 1 of test examination bar sample application zone 13, through sample pad 2, hemofiltration film 3 is to filter haemocyte, blood plasma redissolves and to remove to disturb reagent on enzyme pad 4 (or the pad 5 that flows), finally be penetrated into and flow on pad 5, the going of the endogenous interfering material containing in analyte sample fluid and redissolution disturbs reagent to start disturbance reponse, after the hatching process of 1min left and right, detaching separate layer 12 contacts the reagent layer 6 of promoter region 14 and the mobile pad 5 of sample application zone 13 of test examination bar again, test reaction startup reagent (comprising reaction substrate) and signal fixing on promoter region 14 provide the redissolution such as reagent in analyte sample fluid, and start the test reaction of startup analysis thing, the product generating after test reaction starts provides reagent to carry out signal reaction with signal again, the reflectance value recording according to signal reaction, calculate the content of biology enzyme to be measured.
Embodiment 5:
The dry chemical quantitative test examination bar that a kind of removal is as shown in Figure 6 disturbed, this test examination bar comprises transparent end liner 7, on end liner 7, arrange one and be fixed with the sample application zone 13 (comprising diffusion layer 1, sample pad 2, hemofiltration film 3, enzyme pad 4, mobile pad 5 etc.) of disturbing reagent, sample application zone 13 is directly fixed on the one end on end liner 7; On end liner 7, also arrange one and be fixed with test reaction and start the promoter region 14 that reagent and signal provide reagent, promoter region 14 comprises a reagent layer 6, and reagent layer 6 is directly fixed on the other end of end liner 7.Sample application zone 13 and promoter region 14 are spaced from each other certain distance, make sample application zone 13 and promoter region 14 keep in the normal state noncontact, on end liner 7 in the middle of sample application zone 13 and promoter region 14, be also provided with a crease line 11, under test mode, can, along these crease line 11 turnovers, the promoter region 14 that is positioned at one end be contacted with each other with the sample application zone 13 of the other end.In the present embodiment, the mobile pad 5 of sample application zone 13 is directly fixed on end liner 7, on the pad 5 that flows, is disposed with enzyme pad 4, hemofiltration film 3, sample pad 2 and diffusion layer 1.Go to disturb reagent to be fixed on enzyme pad 4, test reaction starts reagent and signal provides reagent to be fixed on reagent layer 6.
Embodiment 6:
The dry chemical quantitative test examination bar that a kind of removal is as shown in Figure 7 and Figure 8 disturbed, this test examination bar comprises end liner 7, on end liner 7, arrange one and be fixed with the sample application zone 13 (comprising diffusion layer 1, sample pad 2, hemofiltration film 3, enzyme pad 4, mobile pad 5 etc.) of disturbing reagent, sample application zone 13 is directly fixed on end liner 7; On end liner 7, also arranging one is fixed with test reaction and starts the promoter region 14 that reagent and signal provide reagent; promoter region 14 comprises a reagent layer 6 and protective seam 9; protective seam 9 is positioned at the top of reagent layer 6; reagent layer 6 is connected to one end of end liner 7 by an articulated elements 10; this articulated elements 10 has suitable height, and most of region of promoter region 14 reagent layer 6 after horizontal rotary moves can just be contacted with sample application zone 13.In the normal state, promoter region 14 rotations deviate from sample application zone 13 (as shown in Figure 7), keep contactless state with sample application zone 13, can be by itself and sample application zone 13 being contacted with each other around articulated elements 10 rotation promoter regions 14 under test mode.
In the present embodiment, the mobile pad 5 of sample application zone 13 is directly fixed on end liner 7, and one end of the pad 5 that flows is fixed with enzyme pad 4, and the top of enzyme pad 4 is disposed with hemofiltration film 3, sample pad 2 and diffusion layer 1.Enzyme pad 4, hemofiltration film 3, sample pad 2 and diffusion layer 1 are all arranged at a side of the pad 5 that flows, and the space going out more than opposite side is convenient to contact with the reagent layer 6 of promoter region 14.Go to disturb reagent to be fixed on and flow on pad 5, test reaction starts reagent and signal provides reagent to be fixed on reagent layer 6.
Embodiment 7:
A kind of glutamic-pyruvic transaminase quantitative test examination bar of the present invention (GPT test examination bar), this quantitative test examination bar is to be made by the dry chemical quantitative test examination bar of above-described embodiment 5.This test examination bar comprises end liner 7, and end liner 7 adopts transparent polycarbonate (PC) plate, and its thickness is 0.3mm.
The middle part of end liner 7 tops is provided with the pad 5 that flows, and the method for making of the pad 5 that flows in the present embodiment is: the ALANINE PBS solution that contains 0.5M (concentration is 0.1M, pH=7.2) is sprayed on the nitrocellulose filter in approximately 10 μ m apertures, and dry.
One end of mobile pad 5 tops is provided with enzyme pad 4, in the present embodiment, the method for making of enzyme pad 4 is: by the catalatic PBS solution that contains 100KU/L peroxidase, 200KU/L pyruvate oxidase, 100KU/L, (concentration is 0.1M, pH=7.2) be sprayed on the dacron film that 0.09mm is thick, dry.
In sample pad 2, be also provided with diffusion layer 1, the screen cloth form that diffusion layer 1 is made for dacron, aperture is 80 orders.
One end on above-mentioned end liner 7 is also provided with a reagent layer 6, in the present embodiment, the method for making of reagent layer 6 is: in the polycarbonate sheet of the one side frosted that is 0.175mm at thickness by the solution spraying that contains 0.1M α-ketoglutaric acid, 5mM 4-AA (the first developer), 2mM THBHA (chromogen agent), dry.This reagent layer 6 is by bonding mode, to be assembled in one end of end liner 7.As seen from Figure 6, this reagent layer 6 is positioned at a side of the pad 5 that flows, and (not for detection of front) sample application zone 13 and promoter region 14 are spaced from each other certain distance under normal conditions, reagent layer 6 keeps cordless with the pad 5 that flows, owing to being also provided with a crease line 11 on the end liner 7 in the middle of sample application zone 13 and promoter region 14, in carrying out testing process, when analyte sample fluid is penetrated on the pad 5 that flows, reagent layer 6 can be by comprising that any mode of turnover and so on contacts with the pad 5 that flows.
Wherein, the fixing test reaction in promoter region 14 starts reagent and mainly refers to the substrate that can start transamination reaction, in the present embodiment, on reagent layer 6, be fixed with the developer that is applicable to peroxidase and the substrate that can start transamination reaction (being above-mentioned α-ketoglutaric acid).In sample application zone 13, fixing going disturbs reagent mainly to comprise pyruvate oxidase fixing on above-mentioned enzyme pad 4, peroxidase (POD) and hydrogen peroxidase etc.
The glutamic-pyruvic transaminase of a series of variable concentrations is used to Biochemical Analyzer kit definite value (concentration value sees the following form 1), then get respectively the glutamic-pyruvic transaminase solution of 30 μ L variable concentrations as testing sample, in each testing sample, all contain chaff interference pyruvic acid (concentration is 100 μ M), test examination bar and following methods step by above-mentioned the present embodiment are tested the glutamic-pyruvic transaminase content in testing sample: first analyte sample fluid is added drop-wise on the diffusion layer 1 of test examination bar, make flow through successively sample pad 2 and hemofiltration film 3 infiltrating gradually on enzyme pad 4 of analyte sample fluid, catalytic reaction enzyme fixing on enzyme pad 4 (comprises above-mentioned pyruvate oxidase, peroxidase and hydrogen peroxidase etc.) redissolve in analyte sample fluid, the endogenous interfering material pyruvic acid containing in analyte sample fluid reacts with aforesaid catalytic reaction enzyme, analyte sample fluid continues to be penetrated on mobile pad 5, after one section of hatching process (being generally 1min), the reagent layer 6 of test examination bar is contacted with the pad 5 that flows, on reagent layer 6, fixing substrate (α-ketoglutaric acid) and developer etc. redissolve in analyte sample fluid, and start to start transamination reaction, the pyruvic acid generating after transamination reaction starts participates in coupling reaction, the hydrogen peroxide generating after coupling reaction and developer carry out chromogenic reaction, after 2min, according to the reflectance value of chromogenic reagent, also by end-point method (or rate method), determine the content (reflectance value is as shown in table 1 below) of glutamic-pyruvic transaminase.In above-mentioned detection method, before glutamic-pyruvic transaminase is also not activated reaction, endogenous chaff interference pyruvic acid is removed by catalytic reaction.
Get equally the glutamic-pyruvic transaminase solution of above serial variable concentrations as analyte sample fluid, but in each analyte sample fluid, do not contain chaff interference pyruvic acid, above-mentioned test examination bar and the method for testing of same employing tested these analyte sample fluids, test result is as shown in table 1 below, as the comparative example 1 of above-described embodiment.
Get equally the glutamic-pyruvic transaminase solution of above serial variable concentrations as analyte sample fluid, but in each analyte sample fluid, do not contain chaff interference pyruvic acid, same above-mentioned test examination bar (but not containing hydrogen peroxidase on the enzyme pad of test examination bar) and the method for testing of adopting tested these analyte sample fluids, test result is as shown in table 1 below, as the comparative example 2 of above-described embodiment.
The reflectance value of the variable concentrations analyte sample fluid that table 1: embodiment 7 and its comparative example record
According to storehouse Bel's card-Munch (Kubelka-Monk) theorem, reflectivity and testing sample concentration have following linear relationship:
In above formula: R is reflectivity; K, b are constant; C is analyte concentration, and the linear relationship numerical value of above-mentioned comparative example 2 is as shown in table 2 below.
According to the numerical value in following table 2, setting up linear regression equation is y=0.00006x+0.004, R
2=0.986, as shown in figure 10.In formula
x is gpt activity.But, sample with this test examination bar test containing the pyruvic acid (but not containing glutamic-pyruvic transaminase) of 100 μ M, its reflectivity is 85.62%, according to the calculating of regression equation, being equivalent to the testing sample that does not contain glutamic-pyruvic transaminase to detect is the glutamic-pyruvic transaminase solution of concentration 134U/L, is easy to cause false positive results.
Table 2: the reflectance value of the variable concentrations analyte sample fluid that comparative example 2 records and with the linear relationship of concentration value
The reflectivity of comparative example 1 of take is horizontal ordinate, and the reflectivity of the corresponding GPT activity of the present embodiment is ordinate mapping, obtains Figure 11.From table 1 and Figure 11, the testing sample embodiment that contains endogenous interference does not compare with the comparative example 1 of not disturbing containing endogenous, and its reflectance value is very consistent, and it goes interference effect fine as seen.By upper table 1, also can be found out, the noiseless comparative example 1 that contains hydrogen peroxidase (CAT) with do not add catalatic noiseless comparative example 2 and compare, its reflectance value is not significantly improved, and therefore, the hydrogen peroxidase of interpolation does not suppress the colour developing of peroxidase.
For further investigation, continue the following liquid to be measured of GPT test examination bar test with the present embodiment: A liquid (31u/L GPT), B liquid (31u/L GPT, containing 100 μ M pyruvic acid), C liquid (31U/L GPT, containing 150 μ M pyruvic acid), D liquid (31U/LGPT, containing 200 μ M pyruvic acid), result is as shown in table 3 below:
Table 3: test the sample contrast of identical GPT activity, different pyruvic acid concentration
| Solution | A | B | C | D |
| Reflectivity % | 88.09 | 88.18 | 88.15 | 88.06 |
The pyruvic acid of 200 μ M does not have impact substantially on reflectivity as can be seen from Table 3.
Embodiment 8:
A kind of glutamic-oxalacetic transaminease quantitative test examination bar of the present invention (GOT test examination bar), this quantitative test examination bar is to be made by the dry chemical quantitative test examination bar of above-described embodiment 5 equally.Structure, connected mode and the position distribution of each layer of the present embodiment test examination bar are identical with embodiment 7, and, the making material of end liner 7, reagent layer 6, hemofiltration film 3, sample pad 2, diffusion layer 1, making reagent and disposal route are all identical with embodiment 7, are only that the method for making of mobile pad 5 and enzyme pad 4 has a little difference.
In the present embodiment, the method for making of mobile pad 5 is: the L-ASPARTIC ACID PBS solution that contains 0.2M (concentration is 0.1M, pH=7.2) is sprayed on the nitrocellulose filter in approximately 10 μ m apertures, and dry.
In the present embodiment, the method for making of enzyme pad 4 is: the PBS solution that contains 100KU/L peroxidase, 200KU/L pyruvate oxidase (concentration is 0.1M, pH=7.2) is sprayed on the dacron film that 0.09mm is thick, and dry.
The glutamic-oxalacetic transaminease of a series of variable concentrations is used to Biochemical Analyzer kit definite value (concentration value sees the following form 4), then get respectively the glutamic-oxalacetic transaminease solution of 30 μ L variable concentrations as testing sample, in each testing sample, all contain chaff interference pyruvic acid (concentration is 100 μ M), by the test examination bar test glutamic-oxalacetic transaminease content of above-mentioned the present embodiment, method of testing is with embodiment 7, read R1, R2, result is as shown in table 4.
Get equally the glutamic-oxalacetic transaminease solution of above serial variable concentrations as analyte sample fluid, but in each analyte sample fluid, do not contain chaff interference pyruvic acid, above-mentioned test examination bar and the method for testing of same employing tested these analyte sample fluids, test result is as shown in table 4 below, as the comparative example 3 of the present embodiment.
Get equally the glutamic-oxalacetic transaminease solution of above serial variable concentrations as analyte sample fluid, but in each analyte sample fluid, do not contain chaff interference pyruvic acid, same above-mentioned test examination bar (but not containing hydrogen peroxidase on the enzyme pad of test examination bar) and the method for testing of adopting tested these analyte sample fluids, test result is as shown in table 4 below, as the comparative example 4 of the present embodiment.
The reflectance value of the variable concentrations analyte sample fluid that table 4: embodiment 8 and its comparative example record
According to storehouse Bel's card-Munch (K μ belka-Monk) theorem, analyze the linear relationship of comparative example 4 equally, as shown in table 5 below.
According to the numerical value in following table 5, setting up linear regression equation is y=0.00009x+0.003, R
2=0.991, as shown in figure 12.In formula
x is that glutamic-oxalacetic transaminease is active.But, sample with this test examination bar test containing the pyruvic acid (but not containing glutamic-oxalacetic transaminease) of 100 μ M, its reflectivity is 86.75%, according to the calculating of regression equation, being equivalent to the testing sample that does not contain glutamic-oxalacetic transaminease to detect is the glutamic-oxalacetic transaminease solution of concentration 123U/L, is easy to cause false positive results.
Table 5: the reflectance value of the variable concentrations analyte sample fluid that comparative example 4 records and with the linear relationship of concentration value
The reflectivity of comparative example 3 of take is horizontal ordinate, and the reflectivity of the corresponding GOT activity of the present embodiment is ordinate mapping, obtains Figure 13.From table 4 and Figure 13, the testing sample embodiment that contains endogenous interference does not compare with the comparative example 3 of not disturbing containing endogenous, and its reflectance value is very consistent, and it goes interference effect fine as seen.
Similarly, by upper table 4, also can be found out, the noiseless comparative example 3 that contains hydrogen peroxidase (CAT) is compared with not adding catalatic noiseless comparative example 4, and its reflectance value is not significantly improved, therefore, the hydrogen peroxidase of interpolation does not suppress the colour developing of peroxidase.
For further investigation, continue with the GOT test examination bar of the present embodiment, to test following liquid to be measured respectively: A liquid (35u/LGOT), B liquid (35u/L GOT, containing 100 μ M pyruvic acid), C liquid (35u/L GOT, containing 150 μ M pyruvic acid), D liquid (35u/L GOT, containing 200 μ M pyruvic acid), result is as following table 6:
It is active that table 6: embodiment 2 examination bars are tested identical GOT, the sample of different pyruvic acid concentration
| Solution | A | B | C | D |
| Reflectivity % | 88.32 | 88.48 | 88.35 | 88.46 |
The pyruvic acid of 200 μ M does not have impact substantially on reflectivity as can be seen from Table 6.Although above-described embodiment is described with reference to its specific embodiment; but it should be appreciated by those skilled in the art; in the situation that do not depart from spirit of the present invention and the protection domain being defined by the following claims, can carry out to it various modifications of form and details.
Claims (11)
1. a paddy third or glutamic-oxalacetic transaminease quantitative test examination bar, it is characterized in that: described quantitative test examination bar is to be made by dry chemical quantitative test examination bar, described dry chemical quantitative test examination bar comprises end liner, on described end liner, arrange one and be fixed with the sample application zone of disturbing reagent, on described end liner, also arrange one and be fixed with the promoter region that signal provides reagent and test reaction startup reagent, described sample application zone and promoter region are to keep in the normal state noncontact, the mode that can contact with each other under test mode is laid on described test examination bar, test reaction startup reagent fixing on described promoter region mainly refers to the substrate that can start transamination reaction, in described sample application zone, fixing going disturbs reagent to comprise pyruvate oxidase and hydrogen peroxidase, in described promoter region or sample application zone, be also fixed with peroxidase, on described promoter region, fixing signal provides reagent to comprise to be applicable to the chromogenic substrate of described peroxidase, fluorogenic substrate or chemical luminous substrate.
2. paddy third according to claim 1 or glutamic-oxalacetic transaminease quantitative test try bar, it is characterized in that: described sample application zone comprises mobile pad, enzyme pad, hemofiltration film, sample pad and the diffusion layer being fixed on end liner, described enzyme pad is provided with hemofiltration film, described hemofiltration film is provided with sample pad, and described sample pad is provided with diffusion layer.
3. paddy third according to claim 1 and 2 or glutamic-oxalacetic transaminease quantitative test try bar, it is characterized in that: described sample application zone is directly fixed on described end liner, described promoter region is connected on described end liner by a bonding piece, this bonding piece has most of region that enough height make described promoter region and is suspended in described sample application zone top, and this promoter region can be kept in touch by pressing down with described sample application zone.
4. paddy third according to claim 1 and 2 or glutamic-oxalacetic transaminease quantitative test try bar, it is characterized in that: described sample application zone and promoter region are separated by a separate layer, this separate layer is drawable formula connected mode, so that this separate layer is pulled out out described sample application zone and promoter region after test examination bar, can keep in touch.
5. paddy third according to claim 1 and 2 or glutamic-oxalacetic transaminease quantitative test try bar, it is characterized in that: described sample application zone is directly fixed on described end liner, described promoter region is articulated with one end of described end liner by an articulated elements, the rotating shaft of described articulated elements is axially parallel with surface level or vertical, so that promoter region can be kept in touch with described sample application zone after rotating.
6. paddy third according to claim 1 and 2 or glutamic-oxalacetic transaminease quantitative test try bar, it is characterized in that: described sample application zone and promoter region are separately fixed at the zones of different of described end liner the same face, end liner in the middle of described sample application zone and promoter region is provided with a crease line, so that described end liner can make promoter region and sample application zone keep in touch after crease line bending.
7. paddy third according to claim 1 and 2 or glutamic-oxalacetic transaminease quantitative test examination bar, is characterized in that: top, described promoter region is coated with a protective seam, and described protective seam selects transparent polycarbonate or pvc material to make.
8. paddy third according to claim 1 and 2 or glutamic-oxalacetic transaminease quantitative test try bar, it is characterized in that: the concentration of described pyruvate oxidase is 30 KU/L～600KU/L, described catalatic concentration is 1 KU/L～1000KU/L, the concentration of described peroxidase is 30KU/L～600KU/L, and the concentration of described chromogenic substrate, fluorogenic substrate or chemical luminous substrate is 1mM～50mM.
9. paddy third according to claim 1 and 2 or glutamic-oxalacetic transaminease quantitative test examination bar, is characterized in that: the substrate of described startup transamination reaction comprises α-ketoglutaric acid, and the concentration of described α-ketoglutaric acid is 5mM～300mM; The substrate of described startup transamination reaction also comprises for detection of the L-alanine of glutamic-pyruvic transaminase or for detection of the L-L-aminobutanedioic acid of glutamic-oxalacetic transaminease, the concentration of described L-alanine is 0.01M～1M, and the concentration of described L-L-aminobutanedioic acid is 0.01M～1M.
10. one kind is tried with paddy third claimed in claim 1 or glutamic-oxalacetic transaminease quantitative test the method that bar detects paddy third or glutamic-oxalacetic transaminease, comprise the following steps: the sample application zone of first analyte sample fluid being added to described test examination bar, fixing pyruvate oxidase in described sample application zone, hydrogen peroxidase all redissolves in analyte sample fluid, the endogenous interfering material pyruvic acid containing in described analyte sample fluid and pyruvate oxidase and the hydrogen peroxidase in analyte sample fluid start disturbance reponse, after one section of hatching process, make again the promoter region of described test examination bar contact with sample application zone, on promoter region, fixing substrate redissolves in analyte sample fluid, peroxidase fixing in promoter region or sample application zone has also redissolved in analyte sample fluid, now start to start transamination reaction, the pyruvic acid generating after transamination reaction starts participates in coupling reaction, the hydrogen peroxide generating after coupling reaction and described chromogenic substrate carry out chromogenic reaction, according to the reflectance value of chromogenic substrate colour developing and determine the content of paddy third or glutamic-oxalacetic transaminease by end-point method or rate method.
11. 1 kinds of methods that detect paddy third or glutamic-oxalacetic transaminease with paddy third claimed in claim 2 or glutamic-oxalacetic transaminease quantitative test examination bar, comprise the following steps: first analyte sample fluid is added drop-wise on the diffusion layer of described test examination bar sample application zone, make analyte sample fluid flow through successively sample pad and hemofiltration film infiltrate gradually on described enzyme pad, pyruvate oxidase and hydrogen peroxidase fixing in described sample application zone all redissolve in analyte sample fluid, the endogenous interfering material pyruvic acid containing in described analyte sample fluid and pyruvate oxidase and the hydrogen peroxidase in analyte sample fluid start disturbance reponse, after one section of hatching process, make again the promoter region of described test examination bar contact with sample application zone, on promoter region, fixing substrate redissolves in analyte sample fluid, peroxidase fixing in promoter region or sample application zone has also redissolved in analyte sample fluid, now start to start transamination reaction, the pyruvic acid generating after transamination reaction starts participates in coupling reaction, the hydrogen peroxide generating after coupling reaction and described chromogenic substrate carry out chromogenic reaction, according to the reflectance value of chromogenic substrate colour developing and determine the content of paddy third or glutamic-oxalacetic transaminease by end-point method or rate method.
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| CN103776826B (en) * | 2014-01-16 | 2016-05-25 | 无限极（中国）有限公司 | With the test strips of the detection Chinese herbal medicine cadmium content of screening agent and in the purposes detecting in Chinese herbal medicine cadmium content |
| CN106282312A (en) * | 2016-08-19 | 2017-01-04 | 基蛋生物科技股份有限公司 | A kind of dry chemistry bigeminy reagent strip for detecting glutamic oxaloacetic transaminase, GOT and glutamate pyruvate transaminase simultaneously |
| CN109856131B (en) * | 2019-01-22 | 2022-02-18 | 杭州联晟生物科技有限公司 | Test card for simultaneously detecting uric acid, creatinine and urea in sample and preparation method thereof |
| CN109916895A (en) * | 2019-04-12 | 2019-06-21 | 吉林省汇酉生物技术股份有限公司 | A kind of dry chemistry reagent piece and preparation method thereof quantitative determining creatine concentration |
| CN111197071A (en) * | 2020-01-10 | 2020-05-26 | 杭州联晟生物科技有限公司 | Test card for detecting glutamic-pyruvic transaminase by photochemical method and preparation method thereof |
| CN114113247A (en) * | 2020-08-31 | 2022-03-01 | 浙江纳智汇生物科技有限公司 | Electrochemical detection method, detection chip, detection kit and detection system based on enzyme inhibition method |
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