CN102419366A - 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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- CN102419366A CN102419366A CN2011102404608A CN201110240460A CN102419366A CN 102419366 A CN102419366 A CN 102419366A CN 2011102404608 A CN2011102404608 A CN 2011102404608A CN 201110240460 A CN201110240460 A CN 201110240460A CN 102419366 A CN102419366 A CN 102419366A
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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 strip and method of testing, relate in particular to a kind of dry chemical test strip and detection method of transaminase.
Background technology
Chaff interference in the dry chemical test is a lot, the interference of the uric acid in measuring such as redox reaction, ascorbic acid etc., and the pyruvic acid during transaminase detects disturbs, and especially disturbs comparatively serious with the pyruvic acid in the transaminase.
The kind of transaminase is a lot, and is wherein important with glutamic-pyruvic transaminase (GPT/ALT) and glutamic-oxalacetic transaminease (GOT/AST).Glutamic-pyruvic transaminase is claimed ALT again, is the transamination between catalysis glutamic acid and the pyruvic acid; Glutamic-oxalacetic transaminease is claimed the L-aminobutanedioic acid aminopherase again, is the transamination between catalysis glutamic acid and the oxaloacetic acid.Clinically, the mensuration of these two kinds of transaminases in whole blood, serum, blood plasma, the tissue fluid is very important indexs in heart disease, muscle disease, the especially diagnosing hepatism.
Measure the method for above-mentioned two kinds of transaminases clinically; Mainly be to use the method for 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 NADH (NADH), cause the variation of light absorption value at 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 following:
Glutamic-pyruvic transaminase:
Glutamic-oxalacetic transaminease:
The method is consuming time, complicated operation, need more large-scale 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 aspects such as instrument, initial NADH can not be too high, and this makes the test specification of this method be affected, and when test transaminase high value sample, occurs false negative easily.
The dry chemical method has conveniently, flexibly, pollute characteristics such as little, easy and simple to handle.At present, the dry chemical method roughly can be divided into following several types: the electrochemical method (like U.S. Pat 6565738) that with Abbott Laboratories is representative.Fuji, Kodak, Johnson & Johnson are the multilayer diaphragm method (like U.S. Pat 4897347, US5508173, US5462858) of representative.With Luo Shi is the cross flow dry chemical method (like U.S. Pat 4591553, US5508173, US4665023) of representative.Yet; All there is the part defective 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), influence test result than endogenic pyruvic acid.And Fuji is the multilayer diaphragm method of representative, the dry chemical method that Luo Shi is representative, all adopts coupling pyruvate oxidation enzyme process.Adopt the course of reaction of coupling pyruvate oxidation enzyme method following:
Glutamic-pyruvic transaminase is in reaction (1) back coupling
Glutamic-oxalacetic transaminease is in reaction (3) back coupling
According to patent US4665023, the structure of its strip is as shown in Figure 1, and blood sample is added on the diffusion layer 1, and behind hemofiltration film 3 hemofiltrations, blood plasma flows on the pad 5 that flows.1min behind the application of sample presses down transparent protective seam 9, and reagent layer 6 contacts with the pad that flows with enzyme pad 4.The pyruvate oxidase, the peroxidase that are fixed on developer, the zymolyte on the reagent layer 6 and are fixed on the enzyme pad 4 redissolve in sample, thereby produce detectable color signal.
Can find out clearly that from test philosophy there is the interference of endogenous pyruvic acid in this method.The pyruvic acid of color signal is provided, and existing enzymatic produces, and the endogenous pyruvic acid that itself exists in the sample is also arranged.And adopt rate method (i.e. the rate of change of test colour developing), and remove the optical instrument that the endogenous interference needs more complicated, usually adopt optical device such as integrating sphere; And the normal value of transaminase is generally very low; Glutamic-pyruvic transaminase is 5U/L～40U/L, and glutamic-oxalacetic transaminease is 8U/L～40U/L, and the normal reference value of pyruvic acid is higher relatively; It is 65 μ mol/L (being equivalent to the pyruvic acid that the 65U/L transaminase produces at 1min); (in the 2min～3min), the rate of change of chromogenic reagent also can receive the influence of endogenous pyruvic acid to a certain extent, occurs false positive results easily in the short dry chemical test duration.
Summary of the invention
The technical matters that the present invention will solve is the deficiency that overcomes prior art; Provide a kind of measurement result accurately, preparation is simple, testing cost is low, detecting operation is removed interference easily dry chemical quantitative test strip and special quantitative test strip to paddy third or glutamic-oxalacetic transaminease, 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 usefulness or the quantitative test strip detection by quantitative blood two-story valley third of glutamic-oxalacetic transaminease or the method for glutamic-oxalacetic transaminease also are provided.
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 strip of removing interference; Said test strip comprises end liner; Be provided with one on the said end liner and be fixed with the sample application zone of disturbing reagent, also be provided with one on the said end liner and be fixed with the promoter region that signal provides reagent and test reaction to start reagent, said sample application zone and promoter region are to keep noncontact under the state usually, to be laid on the said test strip in the mode that can contact with each other under the test mode.Describedly go to disturb reagent mainly to be meant the reaction reagent that to remove endogenous interfering material in the testing sample; Said signal provides reagent to be meant can and then provide the corresponding reagent of background signal based on the reaction of endogenous interfering material, and described test reaction starts reagent and then comprises preferably reaction reagent, the signal that can start analyte response and produce some biology enzyme reagent that reagent and analytical reactions need etc.
First kind of improvement as the dry chemical quantitative test strip that above-mentioned removal is disturbed; Said sample application zone directly is fixed on the said end liner; Said promoter region is connected on the said end liner through a bonding piece, and this bonding piece has enough height makes most of zone of said promoter region be suspended in said sample application zone top.In this optimized technical scheme, because the promoter region is suspended in the 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 strip, only need the promoter region is pushed downwards, promoter region and sample application zone are kept in touch, to start the relevant detection reaction.
Second kind of improvement as the dry chemical quantitative test strip that above-mentioned removal is disturbed; Said sample application zone and promoter region are separated by a separate layer; This separate layer is set to the connected mode of drawable formula, and said sample application zone and promoter region can keep in touch behind the test strip so that this separate layer detaches out.In this optimized technical scheme,, therefore under normal conditions, exist through the separate layer isolation and with non-contacting state between sample application zone and the promoter region because promoter region and sample application zone are divided on the pros and cons of separate layer; When detecting with this test strip, only need this separate layer is detached out the test strip separately, promoter region and sample application zone are kept in touch, to start the relevant detection reaction.
The third improvement as the dry chemical quantitative test strip that above-mentioned removal is disturbed; Said sample application zone can directly be fixed on the said end liner; Said promoter region then is articulated in an end of said end liner through an articulated elements; The rotating shaft of said articulated elements is axially parallel with surface level or vertical, so that the promoter region can be kept in touch with described sample application zone after rotating.When rotating shaft is axially parallel with surface level, said promoter region be through the upset realization in perpendicular and sample application zone by noncontact to contacting; When rotating shaft is axially vertical with surface level, said promoter region be through the rotation realization in surface level and sample application zone by noncontact to contacting, the height of articulated elements should be suitable at this moment, so that the promoter region can just touch sample application zone after rotating translation.
The 4th kind of improvement as the dry chemical quantitative test strip that above-mentioned removal is disturbed; Said sample application zone and promoter region are separately fixed at the zones of different (each other keep certain distance) of said end liner with one side; End liner in the middle of said sample application zone and the promoter region is provided with a crease line, so that said end liner can make promoter region and sample application zone keep in touch after the crease line bending.
In the dry chemical quantitative test strip that above-mentioned various removals are disturbed; The promoter region can be provided with a reagent layer; Its effect is to start transamination reaction, and preferable material is polycarbonate or nylon, also can material on the reagent layer directly be fixed in the fibrous material as reagent layer; Can set up the protective seam of layer of transparent this moment as required on reagent layer, said protective seam is preferably made with transparent polycarbonate or pvc material.
In the dry chemical quantitative test strip that above-mentioned various removals are disturbed; Structure specific to sample application zone and promoter region; Said sample application zone can be according to the needs of detected sample; Constitute by whole in sample flow pad, diffusion layer, sample pad, hemofiltration film, the enzyme pad or several kinds; Said sample application zone preferably includes and is fixed in the mobile pad on the end liner and is fixed in the enzyme pad on the pad that flows, and said enzyme pad is provided with a kind of or stack in hemofiltration film, sample pad, the diffusion layer wherein multiple is set.For example, can superpose successively on the described enzyme pad hemofiltration film, sample pad and diffusion layer are set, and for example, when detecting serum, just can cancel the hemofiltration rete with this test strip.Said promoter region also can be according to the needs of detected sample, comprise in reagent layer, enzyme pad, the protective seam one or more.
In the dry chemical quantitative test strip that above-mentioned removal is disturbed; Said end liner can adopt (transparent or opaque) macromolecule polymeric material (like tygon, PVC, polystyrene, polyester etc.) to make; But preferably adopt PVC (PVC), polycarbonate, polyamide macromolecule polymeric material to make, the thickness of end liner is preferably 50 μ m～300 μ m.The material that said mobile pad can adopt comprises spun glass, dacron, nitrocellulose filter, PS membrane, poly (ether sulfone) film etc.; Preferred spun glass, nitrocellulose filter, dacron film or the filter paper of adopting is made, and its thickness is preferably 0.04mm～0.2mm.Said enzyme pad preferably adopts spun glass, cellulose filter paper, dacron or filter paper to make.The effect of said hemofiltration film is to filter haemocyte, if sample to be tested is blood plasma, serum then this layer can be set that its material can be various spun glass and the special-purpose hemofiltration film that some producers provide, and preferably adopts the making of asymmetric PS membrane or spun glass.Said sample pad act as the maintenance sample, do not allow sample to overflow (inessential setting), its material can be spun glass, dacron etc., preferably adopts spun glass to make.The effect of said diffusion layer is to make testing sample diafiltration (inessential setting) in lower floor equably; The preferred screen cloth form that adopts dacron or nylon fiber to process; The screen cloth aperture is preferably 40 orders～150 orders; Fibre diameter is preferably 100 μ m～500 μ m, and screen cloth is preferably hydrophilic or hydrophilic treated.
As a total technical conceive; The present invention also provides the method for a kind of usefulness dry chemical quantitative test strip detection of biological enzyme that above-mentioned removal is disturbed; May further comprise the steps: the sample application zone of at first analyte sample fluid being added to said test strip; Fixing going disturbs reagent to redissolve in analyte sample fluid on the said sample application zone, and going in endogenous interfering material that contains in the said analyte sample fluid and the analyte sample fluid disturbs reagent to begin to react, through (keeping 1min to get final product usually) behind one section hatching process; The promoter region of said test strip is contacted with sample application zone; Fixing signal provides reagent and test reaction startup reagent to redissolve in analyte sample fluid on the promoter region, and begins to start the test reaction of analyte, and the product that test reaction starts the back generation provides reagent to carry out signal reaction with said signal again; According to the signal value that signal reaction records, calculate the content of biology enzyme to be measured.In the above-mentioned detection method, the discernible signal that said signal reaction produced comprises and a kind of in color signal, fluorescence signal, the chemiluminescence signal most preferably is color signal that 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 strip; This quantitative test strip is to be processed by above-mentioned dry chemical quantitative test strip; Test reaction startup reagent fixing on the said promoter region mainly is meant the substrate that can start transamination reaction; Fixing going disturbs reagent to comprise pyruvate oxidase and hydrogen peroxidase (CAT) on the said sample application zone, also is fixed with peroxidase (POD) on said promoter region or the sample application zone, and the signal of fixing on the said promoter region provides reagent to comprise to be applicable to chromogenic substrate, fluorogenic substrate or the chemical luminous substrate of said peroxidase.Situation per sample can be added materials such as ascorbic acid oxidase or urate oxidase on the said sample application zone in addition.
As a total technical conceive; The present invention also provides above-mentioned paddy of a kind of usefulness third or glutamic-oxalacetic transaminease quantitative test strip to detect the method (situation of hemofiltration film, sample pad and diffusion layer is not set) of paddy third or glutamic-oxalacetic transaminease; May further comprise the steps: the sample application zone of at first analyte sample fluid being added to said test strip; Pyruvate oxidase, hydrogen peroxidase fixing on the said sample application zone all redissolve in analyte sample fluid; Endogenous interfering material pyruvic acid that contains in the said analyte sample fluid and pyruvate oxidase and the hydrogen peroxidase in the analyte sample fluid begin disturbance reponse, behind one section hatching process, the promoter region of said test strip are contacted with sample application zone; Substrate fixing on the promoter region redissolves in analyte sample fluid; Peroxidase fixing on promoter region or the sample application zone has also redissolved in analyte sample fluid, begins to start transamination reaction this moment, and transamination reaction starts the pyruvic acid of back generation and participates in coupling reaction; Oxydol that generates after the coupling reaction and said chromogenic substrate carry out chromogenic reaction, determine the content of paddy third or glutamic-oxalacetic transaminease according to the reflectance value of chromogenic substrate colour developing and through 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 strip; Said quantitative test strip is had specific sample application zone structure (comprising enzyme pad, hemofiltration film, sample pad and diffusion layer in this sample application zone) dry chemical quantitative test strip and is processed by above-mentioned; Test reaction startup reagent fixing on the said promoter region mainly is meant the substrate that can start transamination reaction; Fixing going disturbs reagent to comprise pyruvate oxidase and hydrogen peroxidase on the said sample application zone; Also be fixed with peroxidase on said promoter region or the sample application zone, fixing signal provides reagent to comprise to be applicable to chromogenic substrate, fluorogenic substrate or the chemical luminous substrate of said peroxidase on the said promoter region; Said enzyme pad is provided with the hemofiltration film, and said hemofiltration film is provided with sample pad, and said sample pad is provided with diffusion layer.Can add buffer system, surfactant, anti-haemolysis reagent (like carbohydrate), salt (like sodium chloride) etc. in diffusion layer, sample pad and the hemofiltration film.
As a total technical conceive; The present invention also provides the method that detects paddy third or glutamic-oxalacetic transaminease with above-mentioned second kind paddy third or glutamic-oxalacetic transaminease quantitative test strip; May further comprise the steps: at first analyte sample fluid is added drop-wise on the diffusion layer of said test strip sample application zone; Make analyte sample fluid flow through successively sample pad and hemofiltration film and soak into gradually on the said enzyme pad; Pyruvate oxidase and hydrogen peroxidase fixing on the said sample application zone all redissolve in analyte sample fluid; Endogenous interfering material pyruvic acid that contains in the said analyte sample fluid and pyruvate oxidase and the hydrogen peroxidase in the analyte sample fluid begin disturbance reponse, behind one section hatching process, the promoter region of said test strip are contacted with sample application zone; Substrate fixing on the promoter region redissolves in analyte sample fluid; Peroxidase fixing on promoter region or the sample application zone has also redissolved in analyte sample fluid, begins to start transamination reaction this moment, and transamination reaction starts the pyruvic acid of back generation and participates in coupling reaction; Oxydol that generates after the coupling reaction and said chromogenic substrate carry out chromogenic reaction, determine the content of paddy third or glutamic-oxalacetic transaminease according to the reflectance value of chromogenic substrate colour developing and through end-point method or rate method.
In above-mentioned paddy third or the glutamic-oxalacetic transaminease quantitative test strip, the concentration of said pyruvate oxidase is preferably 30KU/L～300KU/L.In above-mentioned paddy third or the glutamic-oxalacetic transaminease quantitative test strip, the concentration of said peroxidase is preferably 30KU/L～200KU/L.In above-mentioned paddy third or the glutamic-oxalacetic transaminease quantitative test strip, said catalatic concentration is 1KU/L～200KU/L.In above-mentioned paddy third or the glutamic-oxalacetic transaminease quantitative test strip, the concentration of said chromogenic substrate, fluorogenic substrate or chemical luminous substrate is preferably 1mM～50mM.In the technique scheme, selecting for use of developer can be confirmed (american documentation literatures such as existing US4591553, US5274095, US5162200, US4666023 are mentioned) according to actual needs.If developer system includes only a kind of component, then this component is fixed on the reagent layer of promoter region and gets final product, if developer system comprises various ingredients, then has at least a kind of component to be fixed on the reagent layer of promoter region.
In above-mentioned paddy third or the glutamic-oxalacetic transaminease quantitative test strip, the substrate of said startup transamination reaction preferably includes KG, and the concentration of said KG is preferably 5mM～300mM; The substrate of said startup transamination reaction also preferably includes the L-L-aminobutanedioic acid that is used to detect the L-alanine of glutamic-pyruvic transaminase or is used to detect glutamic-oxalacetic transaminease.In fact, comprised KG, also can be fixed on the sample application zone such as materials such as L-alanine, L-L-aminobutanedioic acids so if be fixed in the substrate of the startup transamination reaction of promoter region.
As do not have especially and mention; The concentration of adhering to material among the present invention on the strip all is meant the concentration that when this strip of preparation, is used for this substance solution that wetting carrier film selects for use, and for example above-mentioned chromogenic agent promptly is meant the concentration that when the preparation strip, is used for this chromogenic reagent solution that wetting carrier film selects for use.
In sum; Reaction system can comprise that (concentration is preferably 30KU/L～300KU/L), peroxidase, and (concentration is preferably 30KU/L～200KU/L), transaminase activator, diphosphothiamine (TPP for buffer system, transamination reaction substrate, anti-haemolysis reagent, pyruvate oxidase in the test strip of the present invention; Concentration is preferably 0.4mg/ml～1mg/m1), flavin adenine dinucleotide (FAD), and (FAD, concentration is preferably 0.1mg/ml～0.2mg/m1), surfactant, developer, pyruvic acid activator (like Mg
2+, Mn
2+, be preferably the Mg of 0.01M～0.1M
2+), paddy third, glutamic-oxalacetic transaminease activator (being preferably 5 ' of 1mM～500mM-phosphopyridoxal pyridoxal phosphate), stabilizing agent etc.Wherein, the activity that comprises selection, concentration and the enzyme of each non-key component of buffer system all can be confirmed (can with reference to american documentation literatures such as US4271265, US4666832, US4591553) according to prior art by those skilled in the art voluntarily.Materials such as TPP, FAD, surfactant, activator all can be fixed on the sample application zone.
Above-mentioned paddy third of the present invention or glutamic-oxalacetic transaminease quantitative test strip and detection method are mainly based on following principle: on the basis of coupling pyruvate oxidase; Can remove the interference of endogenous material pyruvic acid very effectively through adding hydrogen peroxidase again, because the H that endogenous interfering material pyruvic acid produces
2O
2(referring to above-mentioned reaction (5)) are all got rid of through reactions (8) by hydrogen peroxidase, therefore participate in the H of follow-up chromogenic reaction
2O
2(referring to above-mentioned reaction (6)) all are that the product by transamination reaction is transformed.
Compared with prior art; The invention has the advantages that: through adopt test strip of the present invention and detection method can be more accurately, easily glutamic-pyruvic transaminase or glutamic-oxalacetic transaminease are carried out qualitative detection and quantitative test; And test strip of the present invention is simple in structure, is convenient to make, and detection method is easy to operate, quick; Need not newly-increased other instruments or equipment; Meet the requirement of the diagnosis (POCT) of bedside, be fit to uses such as hospital emergency, ward, user oneself, community hospital, have broad application prospects.
Description of drawings
Fig. 1 is the structural representation of dry chemical quantitative test strip in the prior art.
The structural representation of the dry chemical quantitative test strip that Fig. 2 provides for the embodiment of the invention 1 goes to disturb.
The structural representation of the dry chemical quantitative test strip that Fig. 3 provides for the embodiment of the invention 2 goes to disturb.
The structural representation of the dry chemical quantitative test strip that Fig. 4 provides for the embodiment of the invention 3 goes to disturb.
The structural representation of the dry chemical quantitative test strip that Fig. 5 provides for the embodiment of the invention 4 goes to disturb.
The structural representation of the dry chemical quantitative test strip that Fig. 6 provides for the embodiment of the invention 5 goes to disturb.
The structural representation (overlooking) of the dry chemical quantitative test strip that Fig. 7 provides for the embodiment of the invention 6 goes to disturb.
The structural representation (main looking) of the dry chemical quantitative test strip that Fig. 8 provides for the embodiment of the invention 6 goes to disturb.
Another variation of the dry chemical quantitative test strip that Fig. 9 provides for the embodiment of the invention 2 goes to disturb.
The linear regression equation that Figure 10 sets up when carrying out quantitative test for the 2 pairs of glutamic-pyruvic transaminase of Comparative Examples that provide in the embodiment of the invention 7.
Figure 11 is the comparison diagram of reflectance value in the embodiment of the invention 7 and the Comparative Examples 1.
The linear regression equation that Figure 12 sets up when glutamic-oxalacetic transaminease being carried out quantitative test in the Comparative Examples 4 of the present invention.
Figure 13 is the comparison diagram of reflectance value in the embodiment of the invention 8 and the Comparative Examples 3.
Marginal data:
1, diffusion layer; 2, sample pad; 3, hemofiltration film; 4, enzyme pad; 5, mobile 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 present invention is further described.
Embodiment 1:
The dry chemical quantitative test strip that a kind of removal as shown in Figure 2 is disturbed; This test strip comprises end liner 7; Be provided with one on the end liner 7 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 directly is fixed on the end liner 7; Also being provided with one on the end liner 7 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 an end of end liner 7 through a bonding piece 8, and this bonding piece 8 has enough height makes most of zone of reagent layer 6 be suspended in the top of sample application zone 13.Sample application zone 13 keeps noncontact with promoter region 14 under common state, under test mode, can itself and sample application zone 13 (being specially the mobile pad 5 of sample application zone 13) contacted with each other through downward started by press district 14 (reagent layer 6).
In the present embodiment; Sample application zone 13 comprises the mobile pad 5 that is fixed on the end liner 7; Flow and fill up an 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 through bonding piece 8, and finally is fixed on the end liner 7.The reagent layer 6 of promoter region 14 specifically is arranged on mobile pad 5 tops of sample application zone 13.Go to disturb immobilization of reagents on the enzyme pad 4 or the pad 5 that flows, test reaction starts reagent and signal provides immobilization of reagents on reagent layer 6.
The method (being fit to whole blood test, serum, blood plasma etc.) of the dry chemical quantitative test strip detection of biological enzyme that the above-mentioned removal of a kind of usefulness is disturbed; May further comprise the steps: at first analyte sample fluid (as: whole blood) is added on the diffusion layer 1 of test strip sample application zone 13; Process sample pad 2, hemofiltration film 3 are to filter haemocyte; Blood plasma redissolves and to remove to disturb reagent on the enzyme pad 4 or the pad 5 that flows; Being penetrated at last on the pad 5 that flows, the going of the endogenous interfering material that contains in the analyte sample fluid and redissolution disturbs reagent to begin disturbance reponse, pass through the hatching process about 1min after; The reagent layer 6 of the promoter region 14 of test strip is contacted with the mobile pad 5 of sample application zone 13; Fixing test reaction starts reagent (comprising reaction substrate) and signal on the promoter region 14 provides reagent etc. to redissolve in analyte sample fluid, and begins to start the test reaction of analyte, and the product that test reaction starts the back generation provides reagent to carry out signal reaction with signal again; According to the reflectance value that signal reaction records, calculate the content of biology enzyme to be measured.
Embodiment 2:
The dry chemical quantitative test strip that a kind of removal as shown in Figure 3 is disturbed; This test strip comprises end liner 7; Be provided with one on the end liner 7 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 directly is fixed on the end liner 7; Also being provided with one on the end liner 7 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 an end of end liner 7 through a bonding piece 8, and this bonding piece 8 has enough height makes most of zone of reagent layer 6 be suspended in the top of sample application zone 13.Sample application zone 13 keeps noncontact with promoter region 14 under common state, under test mode, can itself and sample application zone 13 (specifically referring to the enzyme pad 4 in the sample application zone 13) contacted with each other through downward started by press district 14 (reagent layer 6).
In the present embodiment; Sample application zone 13 comprises the mobile pad 5 that is fixed on the end liner 7; One end (near an end of promoter region 14) of mobile pad 5 is fixed with enzyme pad 4, and the other end is fixed with hemofiltration film 3, and 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 through bonding piece 8, and finally is fixed on the end liner 7.The reagent layer 6 of promoter region 14 specifically is arranged on enzyme pad 4 tops of sample application zone 13.Go to disturb immobilization of reagents on the enzyme pad 4 or the pad 5 that flows, test reaction starts reagent and signal provides immobilization of reagents on reagent layer 6.
Therefore; Than embodiment 1; Embodiment 2 has done adjustment and optimization with the position of enzyme pad 4, is about to enzyme pad 4 and is fixed on pad 5 ends near promoter region 14 that flow, when detecting; Through downward started by press district 14 it is contacted earlier with enzyme pad 4, the deviation that the difference that 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, and is as shown in Figure 9, makes its part that becomes promoter region 14, goes to disturb this moment reagent should be fixed on the pad 5 that flows.
Embodiment 3:
The dry chemical quantitative test strip that a kind of removal as shown in Figure 4 is disturbed, than the test strip that embodiment 1 provides, the test strip of present embodiment is not established enzyme pad 4, and remaining structure and application mode are identical with embodiment 1.Owing to do not establish enzyme pad 4, the material (for example various catalytic reaction enzyme) that originally was fixed on the enzyme pad 4 can directly be fixed on other structures (pad 5 for example flows) of sample application zone 13.
Embodiment 4:
The dry chemical quantitative test strip that a kind of removal as shown in Figure 5 is disturbed; This test strip comprises end liner 7; Be provided with one on the end liner 7 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 directly is fixed on the end liner 7; Also be provided with one on the end liner 7 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 also directly is fixed on the end liner 7, and sample application zone 13 is passed through a separate layer 12 with promoter region 14 and isolated each other.Separate layer 12 is set to the connected mode of drawable formula, and sample application zone 13 is keeping noncontact under the state usually with promoter region 14, under test mode, can the sample application zone 13 that be positioned at its positive and negative kept in touch mutually with promoter region 14 through detaching separate layer 12.
In the present embodiment; Mobile pad 5 parts of sample application zone 13 are directly fixed on the end liner 7; A part then is positioned at separate layer 12 tops, and the end top 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 immobilization of reagents on enzyme pad 4, test reaction starts reagent and signal provides immobilization of reagents on reagent layer 6.
Method of operating when the test strip of present embodiment is used to detect is similar with embodiment 1; Promptly at first analyte sample fluid (whole blood) is added on the diffusion layer 1 of test strip sample application zone 13; Filtering haemocyte, blood plasma redissolves and removes to disturb reagent on the enzyme pad 4 (or the pad 5 that flows), being penetrated at last on the pad 5 that flows through sample pad 2, hemofiltration film 3; The going of endogenous interfering material that contains in the analyte sample fluid and redissolution disturbs reagent to begin disturbance reponse; Behind the hatching process about 1min, detach separate layer 12 again the reagent layer 6 of the promoter region 14 of test strip is contacted with the mobile pad 5 of sample application zone 13, fixing test reaction startup reagent (comprising reaction substrate) and signal provide reagent etc. to redissolve in analyte sample fluid on the promoter region 14; And begin to start the test reaction of analyte; The product that test reaction starts the back generation provides reagent to carry out signal reaction with signal again, according to the reflectance value that signal reaction records, calculates the content of biology enzyme to be measured.
Embodiment 5:
The dry chemical quantitative test strip that a kind of removal as shown in Figure 6 is disturbed; This test strip comprises transparent end liner 7; Be provided with one on the end liner 7 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 directly is fixed on the end on the end liner 7; Also be provided with one on the end liner 7 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 directly is fixed on the other end of end liner 7.Sample application zone 13 is spaced from each other certain distance with promoter region 14; Make sample application zone 13 and promoter region 14 keep noncontact under the state usually; Also be provided with a crease line 11 on the end liner 7 in the middle of sample application zone 13 and the promoter region 14; Can be under test mode along 11 turnovers of this crease line, the promoter region 14 that is positioned at an end and the sample application zone 13 of the other end are contacted with each other.In the present embodiment, the mobile pad 5 of sample application zone 13 is directly fixed on the end liner 7, is disposed with enzyme pad 4, hemofiltration film 3, sample pad 2 and diffusion layer 1 on the pad 5 that flows.Go to disturb immobilization of reagents on enzyme pad 4, test reaction starts reagent and signal provides immobilization of reagents on reagent layer 6.
Embodiment 6:
A kind of dry chemical quantitative test strip that disturbs like Fig. 7 and removal shown in Figure 8; This test strip comprises end liner 7; Be provided with one on the end liner 7 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 directly is fixed on the end liner 7; Also being provided with one on the end liner 7 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 an end of end liner 7 through an articulated elements 10, and this articulated elements 10 has suitable height, make promoter region 14 through level revolve move after most of zone of reagent layer 6 can just contact with sample application zone 13.Under common state, promoter region 14 rotations deviate from sample application zone 13 (as shown in Figure 7), keep contactless states with sample application zone 13, can be through around articulated elements 10 rotation promoter regions 14 itself and sample application zone 13 being contacted with each other under test mode.
In the present embodiment, the mobile pad 5 of sample application zone 13 is directly fixed on the end liner 7, and an 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 all are arranged at a side of the pad 5 that flows, and the surplus space that goes out of opposite side is convenient to contact with the reagent layer 6 of promoter region 14.Go to disturb immobilization of reagents on the pad 5 that flows, test reaction starts reagent and signal provides immobilization of reagents on reagent layer 6.
Embodiment 7:
A kind of glutamic-pyruvic transaminase quantitative test strip of the present invention (GPT tests strip), this quantitative test strip are to be processed by the dry chemical quantitative test strip of the foregoing description 5.This test strip 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: (concentration is 0.1M, pH=7.2) is sprayed on the nitrocellulose filter in about 10 μ m apertures drying will to contain the L-alanine PBS solution of 0.5M.
The end that fills up 5 tops that flows is provided with enzyme pad 4; The method for making of enzyme pad 4 is in the present embodiment: (concentration is 0.1M will to contain the catalatic PBS solution of 100KU/L peroxidase, 200KU/L pyruvate oxidase, 100KU/L; PH=7.2) be sprayed on the thick dacron film of 0.09mm drying.
Also be provided with diffusion layer 1 on the sample pad 2, the screen cloth form that diffusion layer 1 is processed for dacron, the aperture is 80 orders.
End on the above-mentioned end liner 7 also is provided with a reagent layer 6; The method for making of reagent layer 6 is in the present embodiment: the solution spraying that will contain 0.1M KG, 5mM 4-amino-antipyrine (first developer), 2mM THBHA (chromogen agent) is on the polycarbonate sheet of single face frosted of 0.175mm at thickness, drying.This reagent layer 6 is ends that are assembled in end liner 7 through bonding mode.Visible by Fig. 6; This reagent layer 6 is positioned at a side of the pad 5 that flows, and (before promptly not being used for detecting) sample application zone 13 is spaced from each other certain distance with promoter region 14 under normal conditions, and reagent layer 6 keeps the noncontact mode with the pad 5 that flows; Owing to also be provided with a crease line 11 on the end liner 7 in the middle of sample application zone 13 and the promoter region 14; In carrying out testing process, when being penetrated into to flow, filled up for 5 last times analyte sample fluid, and reagent layer 6 can contact with the pad 5 that flows through any way that comprises turnover and so on.
Wherein, Promoter region 14 fixing test reactions start reagent and mainly are meant the substrate that can start transamination reaction; In present embodiment, be fixed with developer that is applicable to peroxidase and the substrate (being above-mentioned KG) that can start transamination reaction on the reagent layer 6.Fixing going disturbs reagent mainly to comprise pyruvate oxidase, peroxidase (POD) and hydrogen peroxidase etc. fixing on the above-mentioned enzyme pad 4 on the sample application zone 13.
The glutamic-pyruvic transaminase of a series of variable concentrations is used Biochemical Analyzer kit definite value (concentration value sees the following form 1); The glutamic-pyruvic transaminase solution of getting 30 μ L variable concentrations then respectively is as testing sample; All contain chaff interference pyruvic acid (concentration is 100 μ M) in each testing sample; Test strip and following method step through above-mentioned present embodiment are tested the glutamic-pyruvic transaminase content in the testing sample: analyte sample fluid at first is added drop-wise on the diffusion layer 1 of test strip; Make analyte sample fluid flow through sample pad 2 successively with hemofiltration film 3 and soak into gradually on the enzyme pad 4; Catalytic reaction enzyme (comprising above-mentioned pyruvate oxidase, peroxidase and hydrogen peroxidase etc.) fixing on the enzyme pad 4 redissolves in analyte sample fluid; Endogenous interfering material pyruvic acid that contains in the analyte sample fluid and aforesaid catalytic reaction enzyme react, and analyte sample fluid continues to be penetrated on the mobile pad 5, behind one section hatching process (being generally 1min); The reagent layer 6 of test strip is contacted with the pad 5 that flows; Substrate of fixing on the reagent layer 6 (KG) and developer etc. redissolve in analyte sample fluid, and begin to start transamination reaction, and transamination reaction starts the pyruvic acid participation coupling reaction that the back generates; Oxydol that generates after the coupling reaction and developer carry out chromogenic reaction, determine the content (reflectance value is as shown in table 1 below) of glutamic-pyruvic transaminase behind the 2min according to the reflectance value of chromogenic reagent and through end-point method (or rate method).In above-mentioned detection method, glutamic-pyruvic transaminase also is not activated before the reaction, and endogenous chaff interference pyruvic acid is removed through catalytic reaction.
The glutamic-pyruvic transaminase solution of getting above serial variable concentrations equally is as analyte sample fluid; But do not contain the chaff interference pyruvic acid in each analyte sample fluid; Above-mentioned test strip 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 Examples 1 of the foregoing description.
The glutamic-pyruvic transaminase solution of getting above serial variable concentrations equally is as analyte sample fluid; But do not contain the chaff interference pyruvic acid in each analyte sample fluid; Same above-mentioned test strip (but not containing hydrogen peroxidase on the enzyme pad of test strip) and the method for testing of adopting tested these analyte sample fluids; Test result is as shown in table 1 below, as the Comparative Examples 2 of the foregoing description.
The reflectance value of the variable concentrations analyte sample fluid that table 1: embodiment 7 and its Comparative Examples record
According to storehouse Bel's card-Munch (Kubelka-Monk) theorem, reflectivity and testing sample concentration have following linear relationship:
In the following formula: R is a reflectivity; K, b are constant; C is an analyte concentration, and the linear relationship numerical value of above-mentioned Comparative Examples 2 is as shown in table 2 below.
Setting up linear regression equation according to the numerical value in the following table 2 is y=0.00006x+0.004, R
2=0.986, shown in figure 10.
x is a gpt activity in the formula.But; The sample that contains the pyruvic acid (but not containing glutamic-pyruvic transaminase) of 100 μ M with this test strip test; Its reflectivity is 85.62%; According to the calculating of regression equation, be equivalent to the testing sample detection that does not contain glutamic-pyruvic transaminase is the glutamic-pyruvic transaminase solution of concentration 134U/L, be easy to cause false positive results.
Table 2: the reflectance value of the variable concentrations analyte sample fluid that Comparative Examples 2 records and with the linear relationship of concentration value
Reflectivity with Comparative Examples 1 is a horizontal ordinate, and the active reflectivity of the corresponding GPT of present embodiment is the ordinate mapping, gets Figure 11.Visible by table 1 and Figure 11, the testing sample embodiment that contains the endogenous interference compares with the Comparative Examples 1 that does not contain the endogenous interference, and its reflectance value is very consistent, and visible its goes interference effect fine.Can be found out also that by last table 1 the noiseless Comparative Examples 1 that contains hydrogen peroxidase (CAT) is compared with not adding catalatic noiseless Comparative Examples 2, its reflectance value does not obviously improve, and therefore, the hydrogen peroxidase of interpolation does not suppress the colour developing of peroxidase.
For further investigation; Continue with liquid to be measured below the GPT test strip test of present embodiment: A liquid (31u/L GPT), B liquid (31u/L GPT; Contain 100 μ M pyruvic acid), C liquid (31U/L GPT; Contain 150 μ M pyruvic acid), D liquid (31U/LGPT contains 200 μ M pyruvic acid), the 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 that can find out 200 μ M from last table 3 does not have influence basically to reflectivity.
Embodiment 8:
A kind of glutamic-oxalacetic transaminease quantitative test strip of the present invention (GOT tests strip), this quantitative test strip are to be processed by the dry chemical quantitative test strip of the foregoing description 5 equally.Structure, connected mode and the position distribution of each layer of present embodiment test strip are identical with embodiment 7; And; The manufacturing materials 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, only are that the method for making of mobile pad 5 and enzyme pad 4 has a little difference.
The method for making of pad 5 of flowing in the present embodiment is: (concentration is 0.1M, pH=7.2) is sprayed on the nitrocellulose filter in about 10 μ m apertures drying will to contain the L-L-aminobutanedioic acid PBS solution of 0.2M.
The method for making of enzyme pad 4 is in the present embodiment: (concentration is 0.1M, pH=7.2) is sprayed on the thick dacron film of 0.09mm drying will to contain the PBS solution of 100KU/L peroxidase, 200KU/L pyruvate oxidase.
The glutamic-oxalacetic transaminease of a series of variable concentrations is used Biochemical Analyzer kit definite value (concentration value sees the following form 4); The glutamic-oxalacetic transaminease solution of getting 30 μ L variable concentrations then respectively is as testing sample; All contain chaff interference pyruvic acid (concentration is 100 μ M) in each testing sample, through the test strip test glutamic-oxalacetic transaminease content of above-mentioned present embodiment, method of testing is with embodiment 7; Read R1, R2, the result is as shown in table 4.
The glutamic-oxalacetic transaminease solution of getting above serial variable concentrations equally is as analyte sample fluid; But do not contain the chaff interference pyruvic acid in each analyte sample fluid; Above-mentioned test strip 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 Examples 3 of present embodiment.
The glutamic-oxalacetic transaminease solution of getting above serial variable concentrations equally is as analyte sample fluid; But do not contain the chaff interference pyruvic acid in each analyte sample fluid; Same above-mentioned test strip (but not containing hydrogen peroxidase on the enzyme pad of test strip) and the method for testing of adopting tested these analyte sample fluids; Test result is as shown in table 4 below, as the Comparative Examples 4 of present embodiment.
The reflectance value of the variable concentrations analyte sample fluid that table 4: embodiment 8 and its Comparative Examples record
According to storehouse Bel's card-Munch (K μ belka-Monk) theorem, analyze the linear relationship of Comparative Examples 4 equally, as shown in table 5 below.
Setting up linear regression equation according to the numerical value in the following table 5 is y=0.00009x+0.003, R
2=0.991, shown in figure 12.
x is that glutamic-oxalacetic transaminease is active in the formula.But; The sample that contains the pyruvic acid (but not containing glutamic-oxalacetic transaminease) of 100 μ M with this test strip test; Its reflectivity is 86.75%; According to the calculating of regression equation, be equivalent to the testing sample detection that does not contain glutamic-oxalacetic transaminease is the glutamic-oxalacetic transaminease solution of concentration 123U/L, be easy to cause false positive results.
Table 5: the reflectance value of the variable concentrations analyte sample fluid that Comparative Examples 4 records and with the linear relationship of concentration value
Reflectivity with Comparative Examples 3 is a horizontal ordinate, and the active reflectivity of the corresponding GOT of present embodiment is the ordinate mapping, gets Figure 13.Visible by table 4 and Figure 13, the testing sample embodiment that contains the endogenous interference compares with the Comparative Examples 3 that does not contain the endogenous interference, and its reflectance value is very consistent, and visible its goes interference effect fine.
Likewise; Can be found out also that by last table 4 the noiseless Comparative Examples 3 that contains hydrogen peroxidase (CAT) is compared with not adding catalatic noiseless Comparative Examples 4, its reflectance value does not obviously improve; Therefore, the hydrogen peroxidase of interpolation does not suppress the colour developing of peroxidase.
For further investigation; Continue respectively with liquid to be measured below the GOT test strip test of present embodiment: A liquid (35u/LGOT), B liquid (35u/L GOT; Contain 100 μ M pyruvic acid), C liquid (35u/L GOT; Contain 150 μ M pyruvic acid), D liquid (35u/L GOT contains 200 μ M pyruvic acid), result such as following table 6:
It is active that table 6: embodiment 2 strips 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 that can find out 200 μ M from last table 6 does not have influence basically to reflectivity.Although the foregoing description is to describe with reference to its specific embodiment; But it should be appreciated by those skilled in the art; Under the situation that does not break away from the spirit of the present invention that is defined by the following claims and protection domain, can carry out the various modifications of form and details to it.
Claims (14)
1. dry chemical quantitative test strip of removing interference; Said test strip comprises end liner; Be provided with one on the said end liner and be fixed with the sample application zone of disturbing reagent; Also be provided with one on the said end liner and be fixed with the promoter region that signal provides reagent and test reaction startup reagent, it is characterized in that: said sample application zone and promoter region are to keep noncontact under the state usually, to be laid on the said test strip in the mode that can contact with each other under the test mode.
2. the dry chemical quantitative test strip that removal according to claim 1 is disturbed; It is characterized in that: said sample application zone directly is fixed on the said end liner; Said promoter region is connected on the said end liner through a bonding piece; This bonding piece has enough height makes most of zone of said promoter region be suspended in said sample application zone top, and this promoter region can be kept in touch through pressing down with said sample application zone.
3. the dry chemical quantitative test strip that removal according to claim 1 is disturbed; It is characterized in that: said sample application zone and promoter region are separated by a separate layer; This separate layer is drawable formula connected mode, and said sample application zone and promoter region can keep in touch behind the test strip so that this separate layer is detached out.
4. the dry chemical quantitative test strip that removal according to claim 1 is disturbed; It is characterized in that: said sample application zone directly is fixed on the said end liner; Said promoter region is articulated in an end of said end liner through an articulated elements; The rotating shaft of said articulated elements is axially parallel with surface level or vertical, so that the promoter region can be kept in touch with described sample application zone after rotating.
5. the dry chemical quantitative test strip that removal according to claim 1 is disturbed; It is characterized in that: said sample application zone and promoter region are separately fixed at the zones of different of said end liner with one side; End liner in the middle of said sample application zone and the promoter region is provided with a crease line, so that said end liner can make promoter region and sample application zone keep in touch after the crease line bending.
6. the dry chemical quantitative test strip that disturbs according to each described removal in the claim 1～5 is characterized in that: top, said promoter region is coated with a protective seam, and said protective seam selects for use transparent polycarbonate or pvc material to make.
7. the dry chemical quantitative test strip that disturbs according to each described removal in the claim 1～5 is characterized in that: said sample application zone comprises that a kind of or stack in the mobile pad that is fixed on the end liner, enzyme pad, hemofiltration film, sample pad, the diffusion layer is provided with wherein multiple.
8. the method for a dry chemical quantitative test strip detection of biological enzyme that disturbs with each described removal in the claim 1～7; May further comprise the steps: the sample application zone of at first analyte sample fluid being added to said test strip; Fixing going disturbs reagent to redissolve in analyte sample fluid on the said sample application zone; Going in endogenous interfering material that contains in the said analyte sample fluid and the analyte sample fluid disturbs reagent to begin to react; Behind one section hatching process, the promoter region of said test strip is contacted with sample application zone, fixing signal provides reagent and test reaction startup reagent to redissolve in analyte sample fluid on the promoter region; And begin to start the test reaction of analyte; The product that test reaction starts the back generation provides reagent to carry out signal reaction with said signal again, according to the signal value that signal reaction records, calculates the content of biology enzyme to be measured; The discernible signal that said signal reaction produced comprises a kind of in color signal, fluorescence signal, the chemiluminescence signal.
9. paddy third or glutamic-oxalacetic transaminease quantitative test strip; It is characterized in that: said quantitative test strip is to be processed by each described dry chemical quantitative test strip in the claim 1～6; Test reaction startup reagent fixing on the said promoter region mainly is meant the substrate that can start transamination reaction; Fixing going disturbs reagent to comprise pyruvate oxidase and hydrogen peroxidase on the said sample application zone; Also be fixed with peroxidase on said promoter region or the sample application zone, fixing signal provides reagent to comprise to be applicable to chromogenic substrate, fluorogenic substrate or the chemical luminous substrate of said peroxidase on the said promoter region.
10. paddy third or glutamic-oxalacetic transaminease quantitative test strip; It is characterized in that: said quantitative test strip is to be processed by the described dry chemical quantitative test of claim 7 strip; Test reaction startup reagent fixing on the said promoter region mainly is meant the substrate that can start transamination reaction; Fixing going disturbs reagent to comprise pyruvate oxidase and hydrogen peroxidase on the said sample application zone; Also be fixed with peroxidase on said promoter region or the sample application zone, fixing signal provides reagent to comprise to be applicable to chromogenic substrate, fluorogenic substrate or the chemical luminous substrate of said peroxidase on the said promoter region; Said enzyme pad is provided with the hemofiltration film, and said hemofiltration film is provided with sample pad, and said sample pad is provided with diffusion layer.
11. according to claim 9 or 10 described paddy third or glutamic-oxalacetic transaminease quantitative test strip; It is characterized in that: the concentration of said pyruvate oxidase is 30KU/L～600KU/L; Said catalatic concentration is 1KU/L～1000KU/L; The concentration of said peroxidase is 30KU/L～600KU/L, and the concentration of said chromogenic substrate, fluorogenic substrate or chemical luminous substrate is 1mM～50mM.
12. according to claim 9 or 10 described paddy third or glutamic-oxalacetic transaminease quantitative test strip, it is characterized in that: the substrate of said startup transamination reaction comprises KG, the concentration of said KG is 5mM～300mM; The substrate of said startup transamination reaction also comprises the L-L-aminobutanedioic acid that is used to detect the L-alanine of glutamic-pyruvic transaminase or is used to detect glutamic-oxalacetic transaminease, and the concentration of said L-alanine is 0.01M～1M, and the concentration of said L-L-aminobutanedioic acid is 0.01M～1M.
13. method that detects paddy third or glutamic-oxalacetic transaminease with described paddy third of claim 9 or glutamic-oxalacetic transaminease quantitative test strip; May further comprise the steps: the sample application zone of at first analyte sample fluid being added to said test strip; Pyruvate oxidase, hydrogen peroxidase fixing on the said sample application zone all redissolve in analyte sample fluid; Endogenous interfering material pyruvic acid that contains in the said analyte sample fluid and pyruvate oxidase and the hydrogen peroxidase in the analyte sample fluid begin disturbance reponse; Behind one section hatching process; The promoter region of said test strip is contacted with sample application zone, and substrate fixing on the promoter region redissolves in analyte sample fluid, and peroxidase fixing on promoter region or the sample application zone has also redissolved in analyte sample fluid; Begin to start transamination reaction this moment; Transamination reaction starts the pyruvic acid participation coupling reaction that the back generates, and oxydol that generates after the coupling reaction and said chromogenic substrate carry out chromogenic reaction, determines the content of paddy third or glutamic-oxalacetic transaminease according to the reflectance value of chromogenic substrate colour developing and through end-point method or rate method.
14. method that detects paddy third or glutamic-oxalacetic transaminease with described paddy third of claim 10 or glutamic-oxalacetic transaminease quantitative test strip; May further comprise the steps: at first analyte sample fluid is added drop-wise on the diffusion layer of said test strip sample application zone; Make analyte sample fluid flow through successively sample pad and hemofiltration film and soak into gradually on the said enzyme pad; Pyruvate oxidase and hydrogen peroxidase fixing on the said sample application zone all redissolve in analyte sample fluid; Endogenous interfering material pyruvic acid that contains in the said analyte sample fluid and pyruvate oxidase and the hydrogen peroxidase in the analyte sample fluid begin disturbance reponse; Behind one section hatching process; The promoter region of said test strip is contacted with sample application zone, and substrate fixing on the promoter region redissolves in analyte sample fluid, and peroxidase fixing on promoter region or the sample application zone has also redissolved in analyte sample fluid; Begin to start transamination reaction this moment; Transamination reaction starts the pyruvic acid participation coupling reaction that the back generates, and oxydol that generates after the coupling reaction and said chromogenic substrate carry out chromogenic reaction, determines the content of paddy third or glutamic-oxalacetic transaminease according to the reflectance value of chromogenic substrate colour developing and through 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 |
| CN109856131A (en) * | 2019-01-22 | 2019-06-07 | 杭州联晟生物科技有限公司 | It is a kind of for detecting the three-in-one test card and preparation method thereof of uric acid in sample, creatinine, urea 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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