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WO2008141351A1 - Procédé permettant de déterminer de manière quantitative des analytes au moyen d'un élément test ainsi qu'un système de test et son utilisation - Google Patents

Procédé permettant de déterminer de manière quantitative des analytes au moyen d'un élément test ainsi qu'un système de test et son utilisation Download PDF

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Publication number
WO2008141351A1
WO2008141351A1 PCT/AT2008/000172 AT2008000172W WO2008141351A1 WO 2008141351 A1 WO2008141351 A1 WO 2008141351A1 AT 2008000172 W AT2008000172 W AT 2008000172W WO 2008141351 A1 WO2008141351 A1 WO 2008141351A1
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WO
WIPO (PCT)
Prior art keywords
analyte
test
reaction
reaction zone
test system
Prior art date
Application number
PCT/AT2008/000172
Other languages
German (de)
English (en)
Inventor
Alexandra Molinelli
Rudolf Krska
Eva Binder
Johann Binder
Original Assignee
Erber Aktiengesellschaft
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
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Application filed by Erber Aktiengesellschaft filed Critical Erber Aktiengesellschaft
Publication of WO2008141351A1 publication Critical patent/WO2008141351A1/fr

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Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G01N33/54366—Apparatus specially adapted for solid-phase testing
    • G01N33/54386—Analytical elements
    • G01N33/54387—Immunochromatographic test strips
    • G01N33/54388—Immunochromatographic test strips based on lateral flow
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/58—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances
    • G01N33/582—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances with fluorescent label
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/58—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances
    • G01N33/585—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances with a particulate label, e.g. coloured latex

Definitions

  • the present invention relates to a method for the quantitative determination of at least one analyte with a test element in which a quantifiable reaction is carried out on an immunochromatographic membrane fixed on a support on which at least one test or reaction zone is applied. Furthermore, the present invention relates to a test system for the quantitative determination of at least one analyte by means of a quantifiable test element, and to a use of a test system for the quantitative determination of at least one analyte by means of a quantifiable test element.
  • a large number of quantitative or semi-quantitative methods are known in the art with which analytes, in particular biological analytes, can be detected quantitatively or semi-quantitatively. Most of these methods are either time consuming or allow only qualitative or semi-quantitative detection of substances contained in a sample or pollutants.
  • a large number of methods are known in which a thin-film technique is carried out in which test strips or test plates are used and an immunochromatographic membrane is fixed on the test plates with which semi-quantitative or qualitative detection of pollutants contained in samples can be carried out ,
  • Such a semi-quantitative method which uses a thin-layer or strip test with conventional lateral flow, can be found, for example, in WO 00/42434 or GB 2 300 914.
  • this device more than two detection zones are provided on a test strip or element, which are defined along the length of the test strip at distances spaced from each other from the start line, and the amount or semi-quantitative amount of the target analyte can be selected from the number Zones showing a positive result after the sample has been run are determined.
  • EP 0 462 376 discloses an experiment using a double read-out system having a receiving side on which a receiving reagent is applied, which is coated with the analyte with respect to the binding of the labeled Conjugate competes. Further, a conjugate receiving side is provided to which a conjugate-receiving agent is fixed. Immobilization of the conjugate at this point is then related to the amount of analyte in the test sample, with a decrease in the detectable conjugate on the receiving side and a corresponding increase in the detectable conjugate on the conjugate receiving side indicating that an increasing amount of analyte is present in the test sample the sample is present.
  • WO 97/09620 describes quantitative and semi-quantitative experiments in which the signal generated by the target analyte in a detection zone is compared to a signal generated in a range of calibration zones to determine if the signal is Amount of analyte present in the sample is above or below the levels equivalent to those determined by the calibration zones. Standard curve data can be used to calculate the amount of analyte in the sample.
  • the present invention now aims to provide a simple and, in particular, rapid method for the quantitative determination of at least one analyte in a sample. Furthermore, the present invention aims, in addition to the quantitative detection of an analyte with a test element, to provide the quantitative detection of a plurality of analytes contained side by side in a sample safely, quickly and reliably.
  • the method according to the invention for the quantitative of at least one analyte with a test element is essentially characterized in that the at least one analyte and one associated with a dye or fluorescence reaction substance causing associated anti-analyte or the analyte and an analyte labeled with a substance causing a color or fluorescence reaction is brought into contact with the immunochromatographic membrane fixed in a defined amount on the support; in that the at least one analyte and the associated labeled anti-analyte or the analyte and the associated labeled analyte are taken up by the immunochromatographic membrane and by the at least one test or reaction zone containing either the at least one analyte or the corresponding anti-analyte, to be flowed; and that the intensity, in particular the color or fluorescence identity of the at least one analyte and / or the associated anti-analyte in the reaction zone is determined.
  • the at least an analyte which may be dissolved or suspended, and either the corresponding anti-analyte labeled with a substance causing a staining or fluorescence reaction or the substance labeled with a staining or fluorescent reaction via an immunochromato graphic membrane is run and is provided on the immunochromatographic membrane, a test or reaction zone in which either the at least one analyte or the corresponding anti-analyte is included succeeds in the course of a competing reaction between either the labeled anti-analyte or effecting a color reaction in the reaction zone on the labeled analyte and its associated non-labeled anti-analyte or analyte, the color or fluorescence intensity being inversely proportional to the amount of analyte present in the sample.
  • the process is carried out such that the substance causing a color or fluorescence reaction is selected from colored particles, such as gold, latex or silica gel, fluorescent substances, magnetic particles and / or carbon.
  • colored particles such as gold, latex or silica gel, fluorescent substances, magnetic particles and / or carbon.
  • the colored or fluorescent reaction substance from colored particles, magnetic parts and / or carbon, it is possible to achieve a precise grading of the intensities of the at least one or more reaction zones, so that a very accurate detection of the amount in the sample, at least one analyte becomes possible.
  • the method according to the invention is preferably carried out so that the contact time or reaction time of the analyte on the immunochromatographic membrane is determined for the quantitative determination of the amount of analyte absorbed by the immunochromatographic membrane.
  • the contact time or turnover time or transit time of the analyte on the set immunochromatographic membrane succeeds in further clarification of the present method, since in addition to defined amounts of reactants used and the reaction time and thus the amount of solution or suspension, which can be recorded on the immunochromatographic membrane is set.
  • the contact time or reaction time By setting the contact time or reaction time to 15 s to 30 min, in particular 1 min to 10 min, it is ensured that an extremely rapid and reproducible, quantitative detection of at least one analyte is possible.
  • a receiving pad which is designed for receiving the suspension or solution of the analyte, may be fixed on the carrier after the immunochromatographic membrane, thereby determining the amount of liquid which has passed over the immunochromatographic membrane, to ensure and in particular to avoid reflux of this solution or suspension with certainty, which reflux the reproducibility of the process and in particular the accuracy of detection in the quantitative range would be adversely affected.
  • an adsorbent pad can also be used for this purpose.
  • the process according to the invention is preferably carried out such that the test or reaction zone (s) is (are) applied by spraying, dropping or masking.
  • the analyte or the anti-analyte interacting or reacting substances from the analyte itself protein conjugates of the at least one analyte, anti-analyte antibodies, fragments of anti-analyte -Anti emotionsn, proteins, aptamers and conjugates of biological polymers are selected, it is possible, a variety of to be examined, biological or organic substances quickly and accurately, ie quantitatively to investigate by means of the method according to the invention.
  • the inventive method is preferably performed so that the color or fluorescence intensity of the reaction zone by comparison with a background is determined.
  • a comparison zone automatically exists in the method according to the invention, which facilitates the intensity adjustment of the reaction zone in such a way that in the test itself a standard is measured against can, is provided.
  • a standard it is possible, for example, to determine the amount of substance contained in an analyte of interest by simply comparing the sample to be examined with an existing color chart or fluorescence intensities.
  • the method according to the invention is preferably carried out so that the color and fluorescence intensity of the reaction zone in comparison with the background with a photometric or fluorometric measuring device, in particular a photometric or fluorometric reflection device Spectrometer or a CCD camera is measured.
  • a photometric or fluorometric measuring device optical illusions, which can happen when compared with the naked eye, can be avoided with certainty.
  • a completely automated method is provided with which the quantitative amount of an analyte to be examined in a sample can be detected quickly and reliably, and the detection time overall can be lowered further.
  • the error rate of the method according to the invention can be further lowered and the accuracy of the detection further increased become.
  • the inventive method is preferably performed in that additionally a control line or control zone is applied and that as a substrate for the control line or control zone, an immobilized anti-species-specific antibody of the anti-analyte antibody is used.
  • a control line or control zone can be ensured that both the immunochromatographic membrane and at least the at least one reaction zone on this membrane are active and not a faulty test system is present and thus the inventive method was carried out correctly and trouble-free.
  • the inventive method is preferably performed so that the at least one analyte and each having at least two anti-Ana - lyten is contacted, wherein at least one of the anti-analyte is labeled with a dye or fluorescence-causing substance, and the analyte and the associated anti-analytes with the defined in a defined amount on the support immunochromatographic membrane in contact to be brought.
  • the present invention further aims at a rapidly and reliably responding test system for the quantitative determination of at least one analyte by means of a quantifiable test element, with which not only fast but also reliable and reproducible analytes can be detected and identified in biological samples.
  • Such a test system essentially characterized in that a support is provided, on which a defined amount of an immunochromatographic membrane is fixed; in that at least one reaction zone is defined on the immunochromatographic membrane and that a photometric or fluorometric measuring instrument is included for determining the amount of analyte passed through the reaction zone of the test element, the labeled analyte and / or the labeled anti-analyte.
  • a photometric or fluorometric measuring device for determining the amount of an analyte or anti-analyte passed through the reaction zone of the test element, a clearer, faster and more reproducible, quantitative approach is achieved Detection of the analyte.
  • a photometric or fluorometric measuring device a photometric or fluorometric reflection device, a spectrometer or a CCD camera is used, fine color differences of photometric reactions or small reflection differences of fluorometrically reacting substances can be clearly and reproducibly be detected.
  • a portable device as a photometric or fluorometric measuring device, as is the case with a further development of the invention, it is not only possible to provide a small-scale test arrangement, but also a test system with which it is possible to work directly on site, i. For example, at the end user to examine substances for the presence of a questionable analyte.
  • the measurement result is displayed directly on a display of the measuring device, in particular a display, an even further simplification of the test system is provided and it is possible in particular to provide a test system available , which does not require any special technical training, but can be applied by anyone at any location.
  • a test system in which a plurality of test strips is used simultaneously and that the plurality of test strips is fixed in a common holder, a quantitative detection of a plurality of simultaneously in a sample to be examined present, various analytes can be provided in any combination.
  • a system can of course be used for a negative proof or multiple negative and positive proofs side by side.
  • test systems can be provided, for example, in which analytes commonly occurring together on one and the same Test element are applied, so that no more separate detection methods must be performed or different test systems must be used side by side for use, but One and the same test system and method provides the quantitative detection, for example, of several analytes to be examined.
  • the plurality of reaction zones side by side i. at a level at which the test element is applied, it is ensured that, in addition to the detection of possibly several analytes contained in a sample, each analyte can be reproducibly detected over the immunochromatographic membrane after one and the same running time and moreover, for example, by applying different lent concentrations of one and the same substance to be detected or their anti-analytes in adjacent reaction zones, the accuracy of detection is further increased by making a clear assignment of the obtained intensity of the reaction possible.
  • the present invention further aims to use a test system for the quantitative determination of at least one analyte by means of a quantifiable test element, which test system is used for the detection of low molecular weight substances, peptides, fragments of antibodies, aptamers, biological polymers, protein conjugates and protein fragments.
  • the test system is used for the quantitative detection of undesired contaminants and drugs as well as chemical residues, in particular toxins, in particular mycotoxins, in particular trichothecenes, such as nivalenol, deoxynivalenol, 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, fusarenone X, T-2 Toxin, HT-2 toxin, fumonisins such as fumonisin B1, B2 or B3, ochratoxins such as ochratoxin A, B, C or D, zearalenones, moniliformin or aflatoxins such as aflatoxin B1, B2 G1 or G2, M1, M2, alkaloids, Drugs, as well as allergens, pesticides, hormones, antibiotics, additives and additives, ingredients / active ingredients, acrylamides, genetically modified organisms and environmental pollutants and residues used.
  • toxins in particular mycotoxins, in particular trichothece
  • FIG. 1 shows the illustration of a test system according to the present invention with a detection zone and a control zone;
  • Figure 2 is a comparison of standard curves in the detection of deoxynivalenol.
  • Fig. 3 is a regression line for the calibration of a deoxynivalenol strip test;
  • FIG. 4 shows the T2 toxin concentration of extracts measured with a T2 test system;
  • FIG. 5 shows another example of a strip test in which a plurality of punctate reaction areas are shown side by side for the detection of fumonisin B1, aflatoxin B1, chloramphenicol and clenbuterol, wherein FIG. 5a shows a test plate without a control line and FIG. 5b shows the same test plate with a control line; and Fig. 6 shows a test plate in which three allergen-specific monoclonal antibodies, namely hazelnut, walnut and almond, were immobilized on an immunochromatographic membrane.
  • Example 1 a test plate or a test strip for the quantitative detection of deoxynivalenol (DON) in wheat or wheat flour in a concentration range of 250 to 1,600 ppb is shown.
  • the test strip which is generally indicated by 1, has at its start end 2 a release pad 3, which dips into a well, not shown, or a vessel containing the analyte to be examined and either a labeled anti-analyte or a labeled analyte.
  • the sample or analyte to be examined runs in the direction of the arrow 4 over an analytical nitrocellulose membrane 5 which, like the release pad, is applied to a carrier, for example a plastic plate.
  • a reaction line 6 consisting of a protein conjugate of the target analyte deoxynivalenol is applied in the present case.
  • the proteins used for the conjugation were here, for example, bovine serum albumin, ovalbumin min or konalbumin.
  • the reaction zone in the present case was applied to the nitrocellulose membrane 5 using a spray device.
  • a control line 7 consisting of a rabbit anti-mouse antibody was applied to confirm the correct test development since mouse antibodies were used in the experiment.
  • an adsorbent pad 8 At the end of the immunochromatographic membrane 5 is provided for the complete absorption of excess, finally an adsorbent pad 8, in which not only the excess solution is taken up, but also the reflux of the solution is prevented with certainty.
  • Absorbenskissen 8 a cotton pad is applied in the present case.
  • the experimental procedure was conducted as follows. Monoclonal anti-DON antibodies were conjugated to colloidal gold and used as a selective uptake reagent.
  • a soil sample was extracted in a ratio of 1: 4 (weight / volume) with distilled water using a mixer for 3 minutes. The sample was allowed to sit for 5 to 10 minutes. The extract was further diluted 1.2 with distilled water and 50 ⁇ l was used for the test. In the case of too high contaminations with the analyte, obvious dilution series are readily possible.
  • a sufficient amount of monoclonal antibody, in the present mouse monoclonal antibody, is used for the coupling reaction with colloidal gold, which has particle sizes of about 40 nm.
  • colloidal gold which has particle sizes of about 40 nm.
  • the calculated amount of antibody was added to 1 ml of distilled water at pH 8.5.
  • the antibody solution was then mixed with a colloidal gold solution in a centrifuge tube and mixed for 90 minutes at room temperature with gentle stirring on a shaking platform.
  • the unreacted residual binding sites on the mouse monoclonal antibody were bound by the addition of 2% BSA or bovine serum albumin and incubation of the mixture for a further 90 min.
  • To separate unbound antibody was centrifuged for 30 min at 8000 G and washed with 30 mL of distilled water at pH 8.5.
  • test extract was thoroughly mixed with the receiving reagent buffer mixture in a microtiter well by repeated pipetting.
  • the strip test was then vertically immersed in the well containing a well-defined amount of solution and uptake reagent for a test period of 3 minutes.
  • the colloidal gold allowed the development of colored red lines after selective retention by the immobilized reagents.
  • the reaction line 6 was applied at 8.0 ⁇ 5 mm from the bottom of the immunochromatographic membrane 5 and had a width of about 1.0 ⁇ 1 mm.
  • Control line 7 had a constant intensity independent of the presence or absence of the target analyte for rapid visual confirmation of the correct test performance and to confirm that the test plate used is active and effective.
  • Control line 7 was applied at about 13 to 14 mm from the bottom of the immunochromatographic membrane 5 and had a distance from the absorbent pad of about 3 mm.
  • a positive sample resulted in a slightly colored line in the reaction zone 6.
  • a negative sample would result in the formation of a clearly visible line in the reaction zone 6, since in the present case a competing reaction between the analyte to be detected and the labeled anti-analyte and the Analyte in the reaction zone 6 took place.
  • the test strip 1 was then developed with a portable reader to scan the reaction zone 6. The test results can be read immediately after the three-minute test development time.
  • FIG. 2 shows the comparison of two standard curves in the range from 0 to 2,000 ppb DON, wherein in particular in the curve 9 a straight region 10 clearly occurs, in which the experiment can be carried out in a quantitatively reproducible manner.
  • the portable reader can perform an evaluation of the experiment immediately and without delay. It is irrelevant to note that when a fluorescent reagent is used instead of colloidal gold, an evaluation can be made immediately using, for example, a naked-eye UV lamp and comparison with test cards.
  • Fig. 3 shows a calibration of a selected working range of the test system using deoxynivalenol.
  • the method used may be matrix-dependent and must be chosen according to the matrix to be investigated.
  • the reaction zone signal is immediately measured at the end of the test period with a reader and the data are processed with available software, which immediately yields a quantitative result.
  • Example 2 which is conducted essentially as Example 1, instead of the quantitative detection of deoxynivalenol, that of a concentration of T2 toxin extract is detected, which is measured with a specific T2 strip test.
  • the test strip for the assay plate is hereby identical to that of Example 1 and as shown in Fig. 1, except that anti-T2 monoclonal antibodies were conjugated to colloidal gold and the labeled anti-analytes in the represent sample to be examined.
  • the test procedure was otherwise carried out as in Example 1, except that the test duration was 5 minutes.
  • the range of activity of the quantitative test for T2 toxin is in the range of 1 to 100 ppb.
  • FIG. 5 a shows a test system in which the immunochromatographic membrane 5 is brought into direct contact with the analyte to be detected or the labeled anti-analyte or the labeled analyte. A release zone is not provided.
  • a plurality of punctiform reaction zones 6 are applied to the immunochromatographic membrane 5, each of the reaction zones containing an analyte-protein conjugate.
  • the test system according to FIG. 5a also has an absorbent pad 8 for receiving excess reagent or excess solution containing the substance to be detected.
  • FIG. 5b shows the analogous immunochromatographic membrane 5, in which in turn reaction zones or test points 6 containing four different analyte-protein conjugates are applied.
  • Fig. 5b has a control line 7 for proving whether the test is positive, which is formed as in Example 1.
  • an absorbent pad 8 for receiving excess analyte solution or suspension is also provided in FIG. 5b in order to avoid a backflow of the analyte into the reaction system. More specifically, Example 3 is executed as follows.
  • Protein conjugates coupled to BSA and / or ovalbumin and conalbumin A, of fumonisin B1 and aflatoxin B1, chloramphenicol and clenbuterol were applied as circular surfaces 6.
  • a rabbit anti-mouse antibody was used to check the correctness of the test procedure.
  • concentrations of the conjugates were used for test optimization in the range of 0.05 mg / ml to 5 mg / ml.
  • a mixture of monoclonal or polyclonal antibodies conjugated to colloidal gold and each with specific affinity for each analyte was used to selectively recognize and bind the target analytes.
  • the intensity of the circular zones was measured with a suitable reader and quantified by an externally created calibration function.
  • the target analytes were in diluted form, on the one hand as a standard dissolved in methanolic solution (solution 1, as shown in Table 1), and on the other hand on a real sample extract from animal feed, as shown in Table 2, tested with the target substances.
  • the results are shown in the form of signal intensities as follows, converted into concentration values by means of the external calibration function.
  • a test was carried out for the simultaneous detection of allergenic hazelnut, walnut and almond proteins.
  • a nitrocellulose analytical membrane 5 was used as the immunochromatographic membrane in combination with a release pad 3.
  • the samples to be examined ran in the direction of arrow 4.
  • On the nitrocellulose membrane 5 were three reaction or test zones 6 of one reagent at the same height from the bottom of the test plate 1 for simultaneous detection of the three allergenic nut proteins of aqueous Extracts from foods believed to contain these nuts are applied.
  • Reaction zones 6 were monoclonal antibodies to the following target proteins, selected from Cor a11 for hazelnut (48 kDa allergen), Jug r2 for walnut (45 kDa allergen) and Pru du4 for almond (14 kDa allergen).
  • the reaction zones 6 had a maximum area of 5 mm ⁇ 5 mm.
  • the applied concentrations ranged from 0.5 to 5 mg / L to bind the available target allergenic protein.
  • a rabbit anti-mouse antibody was used to confirm the correct test development. Control line 7 at 0.75 mg / ml was sprayed 1 ⁇ l / cm onto a contact point delivery platform.
  • Polyclonal antibodies were used as the second antibody in the sandwich experiments and conjugated with colloidal gold for use as a selective uptake reagent.
  • a mixture of uptake reagents for each target allergen was mixed.
  • the sample extract was thoroughly mixed in a well with the uptake-agent buffer mixture in a 1: 1 ratio by repeated pipetting and release.
  • the prepared mixture containing the extract was applied in a metered amount to the release pad 3 of the strip test.
  • the liquid sample was allowed to flow on the membrane for a test period of 10 minutes and the excess was collected on an absorbent pad 8.
  • the colloidal gold allowed the development of colored red zones after selective retention of the allergen proteins by the immobilized reagents.
  • the control line had a constant intensity independent of the presence or absence of the target allergens for rapid, visual confirmation that the test is functioning correctly.
  • a positive sample resulted in the formation of a visible reaction zone.
  • a negative sample resulted in no signal formation on the Reaction zone.
  • the signal intensity on each reaction zone was proportional to the protein concentration.
  • Walnut, almond and hazelnut samples were used to determine the specificity of the antibodies. Roasted hazelnuts, white almonds and untreated walnuts were used. For each chosen nut, 200 grams of ground nuts were mixed with 400 ml of ice cold distilled water, dispersed in a mixer at 6N for about 90 seconds, and passed through a 125 ⁇ m sieve for 30 minutes. The supernatant was again dispersed with 200 ml of ice-cold distilled water and again passed through a sieve. The sieve fractions were lyophilized overnight to total dryness. The resulting powder was stored at -20 0 C until further use.
  • the nut strip test was developed for use with a meter which is used to sense the intensity of the reaction zones 6. Depending on the type of meter, a mean, relative intensity or average pixel intensity in the reaction zone 6 is measured. The selection of the area size of the reaction zone makes it possible to adjust or adapt a possible concentration range of the selected target proteins which are scanned. The intensity of reaction zone 6 may decrease from the bottom to the top of the test zone in the direction of flow, depending on the amount of allergenic proteins in the sample. The results may be expressed in ⁇ g / ml protein extract or mg nut / kg of tested food.
  • Control line 7 was visible within one minute, confirming the correct test development, while 10 to 15 minutes were required for complete signal development at reaction zones 6, depending on the matrix chosen.
  • test results could be read directly after the test development time.

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Abstract

L'invention concerne un procédé et un système de test permettant de déterminer quantitativement au moins un analyte au moyen d'un élément test, selon lequel une transformation quantifiable est réalisée sur une membrane (5) immunochromatographique et fixée à un support, sur laquelle au moins une zone (6) de réaction ou de test est appliquée. L'invention concerne également son utilisation, de sorte qu'au moins l'analyte et un anti-analyte correspondant, marqué par une substance agissant par une réaction colorée ou par fluorescence ou l'analyte et un analyte marqué par un substance agissant par une réaction colorée ou par fluorescence est mis en contact avec la membrane (5) immunochromatographique et appliqueé en quantité définie sur le support, que l'analyte et l'anti-analyte marqué correspondant ou l'analyte et l'analyte marqué correspondant sont absorbés par la membrane (5) immunochromatographique et peuvent s'écouler à travers la zone (6) de réaction ou de test, contenant au moins un analyte ou l'anti-analyte associé; et que l'intensité, en particulier l'intensité de la couleur ou de la fluorescence, dudit analyte et/ou de l'anti-analyte associé est déterminée dans la zone (6) de réaction.
PCT/AT2008/000172 2007-05-21 2008-05-15 Procédé permettant de déterminer de manière quantitative des analytes au moyen d'un élément test ainsi qu'un système de test et son utilisation WO2008141351A1 (fr)

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ATGM324/2007 2007-05-21
AT3242007 2007-05-21

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CN101900728A (zh) * 2010-08-05 2010-12-01 中国农业科学院油料作物研究所 半定量化检测黄曲霉毒素b1的多检测线免疫层析试纸条及其制备方法
CN101930006A (zh) * 2010-08-05 2010-12-29 中国农业科学院油料作物研究所 快速检测黄曲霉毒素总量的高灵敏度免疫层析试纸条及其制备方法
CN102253209A (zh) * 2010-12-21 2011-11-23 中国农业科学院油料作物研究所 黄曲霉毒素m1免疫层析试纸条及其制备方法
CN102279268A (zh) * 2011-07-29 2011-12-14 上海交通大学 同时检测玉米赤霉烯酮与伏马毒素的方法
CN102478577A (zh) * 2010-11-30 2012-05-30 吉林大学 一种检测伏马菌素的化学发光试剂盒及其制备方法
CN102520179A (zh) * 2011-12-07 2012-06-27 上海交通大学 定量检测伏马毒素b1的方法
CN102517291A (zh) * 2011-11-25 2012-06-27 国家纳米技术与工程研究院 伏马毒素b1核酸适体及其应用
CN102520177A (zh) * 2011-12-07 2012-06-27 上海交通大学 定量检测玉米赤霉烯酮的方法
CN102539772A (zh) * 2012-01-06 2012-07-04 上海交通大学 基于免疫磁酶的检测伏马毒素b1的方法
CN102559686A (zh) * 2011-11-25 2012-07-11 国家纳米技术与工程研究院 脱氧雪腐镰刀菌烯醇核酸适体及其应用
CN102590364A (zh) * 2011-01-05 2012-07-18 中国医学科学院药用植物研究所 一种同时检测不同基质中药中伏马毒素b1、b2的方法
CN102955031A (zh) * 2011-08-31 2013-03-06 北京勤邦生物技术有限公司 检测黄曲霉毒素b1药物的酶联免疫试剂盒及其应用
CN103063831A (zh) * 2013-01-15 2013-04-24 国家烟草质量监督检验中心 烟草及烟草制品中黄曲霉毒素的酶联免疫测定方法
RU2480750C1 (ru) * 2011-09-27 2013-04-27 Государственное бюджетное образовательное учреждение высшего профессионального образования "Уральская государственная медицинская академия Министерства здравоохранения и социального развития Российской Федерации" (ГБОУ ВПО УГМА Минздравсоцразвития России) Способ определения индивидуальной дозы аллергена для проведения аллергопробы in vitro
CN103792347A (zh) * 2012-11-05 2014-05-14 江苏维赛科技生物发展有限公司 B类单端孢霉烯族毒素的酶联免疫吸附检测专用测试盒
CN103792365A (zh) * 2012-11-05 2014-05-14 江苏维赛科技生物发展有限公司 串珠镰刀菌素胶体金试纸条的制备及使用方法
CN103792348A (zh) * 2012-11-05 2014-05-14 江苏维赛科技生物发展有限公司 一种b类单端孢霉烯族毒素半定量速测试剂板的制备
CN103792356A (zh) * 2012-11-04 2014-05-14 江苏维赛科技生物发展有限公司 检测伏马毒素的elisa试剂盒的制备及检测方法
CN103808938A (zh) * 2014-03-16 2014-05-21 吉林大学 一种检测玉米赤酶烯酮的方法
CN103808920A (zh) * 2012-11-06 2014-05-21 江苏维赛科技生物发展有限公司 串珠镰刀菌素酶联免疫检测试剂盒
CN104007255A (zh) * 2014-05-16 2014-08-27 中南林业科技大学 一种黄曲霉素b1快速检测试纸条及其制备方法与应用
CN104198711A (zh) * 2014-08-25 2014-12-10 广东省农业科学院动物卫生研究所 一种免疫磁珠及其在检测玉米赤霉烯酮中的应用
CN104914196A (zh) * 2015-06-19 2015-09-16 广西壮族自治区梧州食品药品检验所 食品中黄曲霉毒素的测定方法
CN105067811A (zh) * 2015-07-22 2015-11-18 中国农业大学 基于荧光微球免疫层析法检测t-2毒素的产品及其制备方法
CN106093406A (zh) * 2016-05-31 2016-11-09 东北农业大学 玉米赤霉烯酮和脱氧雪腐镰刀菌烯醇双检试纸条
CN106771213A (zh) * 2016-11-29 2017-05-31 百奥森（江苏）食品安全科技有限公司 一种伏马毒素检测方法
CN109001190A (zh) * 2018-05-29 2018-12-14 中国农业科学院农业质量标准与检测技术研究所 基于比色卡鉴别的黄曲霉毒素吸附剂现场评价试剂装置的制备方法
CN109085336A (zh) * 2018-08-29 2018-12-25 郑州工程技术学院 一种检测伏马菌素b1的免疫层析试纸
CN109709321A (zh) * 2019-01-08 2019-05-03 中国科学院生态环境研究中心 一种酶联核酸适配体微孔板光学分析小分子的方法
CN110488016A (zh) * 2019-08-14 2019-11-22 江南大学 一种玉米赤霉烯酮-呕吐毒素双通道免疫定量试纸条

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CN101930006A (zh) * 2010-08-05 2010-12-29 中国农业科学院油料作物研究所 快速检测黄曲霉毒素总量的高灵敏度免疫层析试纸条及其制备方法
CN101900728A (zh) * 2010-08-05 2010-12-01 中国农业科学院油料作物研究所 半定量化检测黄曲霉毒素b1的多检测线免疫层析试纸条及其制备方法
CN101930006B (zh) * 2010-08-05 2012-08-22 中国农业科学院油料作物研究所 快速检测黄曲霉毒素总量的高灵敏度免疫层析试纸条及其制备方法
CN102478577A (zh) * 2010-11-30 2012-05-30 吉林大学 一种检测伏马菌素的化学发光试剂盒及其制备方法
CN102253209A (zh) * 2010-12-21 2011-11-23 中国农业科学院油料作物研究所 黄曲霉毒素m1免疫层析试纸条及其制备方法
CN102253209B (zh) * 2010-12-21 2013-07-17 中国农业科学院油料作物研究所 黄曲霉毒素m1免疫层析试纸条及其制备方法
CN102590364A (zh) * 2011-01-05 2012-07-18 中国医学科学院药用植物研究所 一种同时检测不同基质中药中伏马毒素b1、b2的方法
CN102279268A (zh) * 2011-07-29 2011-12-14 上海交通大学 同时检测玉米赤霉烯酮与伏马毒素的方法
CN102955031A (zh) * 2011-08-31 2013-03-06 北京勤邦生物技术有限公司 检测黄曲霉毒素b1药物的酶联免疫试剂盒及其应用
CN102955031B (zh) * 2011-08-31 2015-07-01 北京勤邦生物技术有限公司 检测黄曲霉毒素b1药物的酶联免疫试剂盒及其应用
RU2480750C1 (ru) * 2011-09-27 2013-04-27 Государственное бюджетное образовательное учреждение высшего профессионального образования "Уральская государственная медицинская академия Министерства здравоохранения и социального развития Российской Федерации" (ГБОУ ВПО УГМА Минздравсоцразвития России) Способ определения индивидуальной дозы аллергена для проведения аллергопробы in vitro
CN102559686A (zh) * 2011-11-25 2012-07-11 国家纳米技术与工程研究院 脱氧雪腐镰刀菌烯醇核酸适体及其应用
CN102517291A (zh) * 2011-11-25 2012-06-27 国家纳米技术与工程研究院 伏马毒素b1核酸适体及其应用
CN102520177A (zh) * 2011-12-07 2012-06-27 上海交通大学 定量检测玉米赤霉烯酮的方法
CN102520179A (zh) * 2011-12-07 2012-06-27 上海交通大学 定量检测伏马毒素b1的方法
CN102539772A (zh) * 2012-01-06 2012-07-04 上海交通大学 基于免疫磁酶的检测伏马毒素b1的方法
CN102539772B (zh) * 2012-01-06 2014-08-06 上海交通大学 基于免疫磁酶的检测伏马毒素b1的方法
CN103792356A (zh) * 2012-11-04 2014-05-14 江苏维赛科技生物发展有限公司 检测伏马毒素的elisa试剂盒的制备及检测方法
CN103792347A (zh) * 2012-11-05 2014-05-14 江苏维赛科技生物发展有限公司 B类单端孢霉烯族毒素的酶联免疫吸附检测专用测试盒
CN103792348A (zh) * 2012-11-05 2014-05-14 江苏维赛科技生物发展有限公司 一种b类单端孢霉烯族毒素半定量速测试剂板的制备
CN103792365A (zh) * 2012-11-05 2014-05-14 江苏维赛科技生物发展有限公司 串珠镰刀菌素胶体金试纸条的制备及使用方法
CN103808920A (zh) * 2012-11-06 2014-05-21 江苏维赛科技生物发展有限公司 串珠镰刀菌素酶联免疫检测试剂盒
CN103063831A (zh) * 2013-01-15 2013-04-24 国家烟草质量监督检验中心 烟草及烟草制品中黄曲霉毒素的酶联免疫测定方法
CN103808938A (zh) * 2014-03-16 2014-05-21 吉林大学 一种检测玉米赤酶烯酮的方法
CN104007255A (zh) * 2014-05-16 2014-08-27 中南林业科技大学 一种黄曲霉素b1快速检测试纸条及其制备方法与应用
CN104198711A (zh) * 2014-08-25 2014-12-10 广东省农业科学院动物卫生研究所 一种免疫磁珠及其在检测玉米赤霉烯酮中的应用
CN104914196A (zh) * 2015-06-19 2015-09-16 广西壮族自治区梧州食品药品检验所 食品中黄曲霉毒素的测定方法
CN105067811A (zh) * 2015-07-22 2015-11-18 中国农业大学 基于荧光微球免疫层析法检测t-2毒素的产品及其制备方法
CN106093406A (zh) * 2016-05-31 2016-11-09 东北农业大学 玉米赤霉烯酮和脱氧雪腐镰刀菌烯醇双检试纸条
CN106771213A (zh) * 2016-11-29 2017-05-31 百奥森（江苏）食品安全科技有限公司 一种伏马毒素检测方法
CN109001190A (zh) * 2018-05-29 2018-12-14 中国农业科学院农业质量标准与检测技术研究所 基于比色卡鉴别的黄曲霉毒素吸附剂现场评价试剂装置的制备方法
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CN109085336B (zh) * 2018-08-29 2021-03-02 郑州工程技术学院 一种检测伏马菌素b1的免疫层析试纸
CN109709321A (zh) * 2019-01-08 2019-05-03 中国科学院生态环境研究中心 一种酶联核酸适配体微孔板光学分析小分子的方法
CN110488016A (zh) * 2019-08-14 2019-11-22 江南大学 一种玉米赤霉烯酮-呕吐毒素双通道免疫定量试纸条

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