CN115840042B - Immunochromatography test paper box and preparation method and application thereof - Google Patents
Immunochromatography test paper box and preparation method and application thereof Download PDFInfo
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- CN115840042B CN115840042B CN202310162831.8A CN202310162831A CN115840042B CN 115840042 B CN115840042 B CN 115840042B CN 202310162831 A CN202310162831 A CN 202310162831A CN 115840042 B CN115840042 B CN 115840042B
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Abstract
The invention relates to an immunochromatography test paper box and a preparation method and application thereof. The immunochromatography test paper box comprises a test paper strip and a sample diluent containing 0.15-0.25 w/v% of tween, 0.05-0.15 w/v% of PEG, 0.05-0.15 w/v% of surfactant, 0.05-0.15 w/v% of biological preservative and 0.15-0.25 w/v% of sodium sulfide; the sample pad of the test strip is treated by a sample pad treatment liquid containing 0.45-0.55 w/v% BSA, 0.15-0.25 w/v% Tween, 0.05-0.15 w/v% PEG, 0.8-1.5 w/v% HBR, 0.8-1.5 w/v% resin and 0.45-0.55 w/v% RBC. The invention can effectively seal the interference factors in the feces, eliminate the nonspecific interference of animal feces samples in immunochromatography detection, and improve the detection sensitivity and accuracy.
Description
Technical Field
The invention belongs to the technical field of immunochromatography, relates to an immunochromatography test paper box and a preparation method and application thereof, and particularly relates to an immunochromatography test paper box for eliminating non-specific interference of animal faeces samples and a preparation method and application thereof.
Background
In-vitro animal diagnosis reagents need to be used for fecal samples such as feline pestivirus (feline panleukopenia virus), but components such as fecal bile pigment, urobilin, heavy metals, bacterial proteins and shed cell proteins in animal fecal components can influence experimental detection results. Taking bacterial proteins and shed cell proteins as examples, animal feces contain a large number of and various bacteria, such as Badong bacillus and salmonellosis in cat feces, and may also carry parasites, such as toxoplasma, hookworm and the like, and many proteins (such as bacterial proteins and shed cell proteins) contained in the organisms can serve as interfering antibodies, cross-linking capture antibodies and detection antibodies, resulting in high readings in analysis, thus causing problems of false positive results and the like.
The method for processing the sample in the immunochromatography technology comprises the following steps: adding a sample diluent; coating a blocking agent on the NC film to form a blocking line; blocking agents and the like are added to the treatment formulation of the sample pad. Most of sample diluents in the market comprise sugar, salt, sealing agent and the like; the formula of the sample pad is a sealing agent, a surfactant, a buffer solution and the like, for example, the conventional formula of the prior art of the sample pad is BSA, tween, PVP-K30 and the like, the sample pad is dried at 37 ℃ for 15 hours, and the sample pad is taken out and assembled with a reaction membrane and a gold mark pad; this conventional technique cannot realize the false positive problem of in vitro diagnosis and detection of fecal samples for pets. The interference components in the pet feces are as follows: the prior art cannot well seal the interfering substances, such as the urobilin, heavy metals, bacterial proteins, abscisic cell proteins and the like, and the interfering substances are combined with the coating antibody on the reaction membrane during sample addition detection, so that the false positive problem is generated.
In summary, it is very necessary to provide an immunochromatographic test paper cassette, and a preparation method and application thereof.
Disclosure of Invention
In order to solve the technical problems in the prior art, the invention provides an immunochromatography test paper box, and a preparation method and application thereof. According to the immunochromatography test paper box, the sample diluent with a proper formula is matched with the sample pad treatment liquid, so that interference factors in feces can be effectively blocked, nonspecific interference of animal feces samples in immunochromatography detection can be eliminated, and the sensitivity and accuracy of detection are effectively improved.
The present invention provides in a first aspect an immunochromatographic test strip cartridge comprising an immunochromatographic test strip and a sample diluent; the sample diluent comprises 0.15-0.25 w/v% of tween, 0.05-0.15 w/v% of polyethylene glycol, 0.05-0.15 w/v% of surfactant, 0.05-0.15 w/v% of biological preservative and 0.15-0.25 w/v% of sodium sulfide; the sample pad of the immunochromatographic test strip is treated by a sample pad treatment liquid; the sample pad treatment solution contains 0.45-0.55 w/v% of bovine serum albumin, 0.15-0.25 w/v% of tween, 0.05-0.15 w/v% of polyethylene glycol, 0.8-1.5 w/v% of an amphotropic antibody blocker, 0.8-1.5 w/v% of resin and 0.45-0.55 w/v% of a rabbit anti-human polyclonal antibody.
Preferably, the mass ratio of the bovine serum albumin, tween, polyethylene glycol, the amphotropic antibody blocking agent, the resin and the rabbit anti-human polyclonal antibody in the sample pad treatment solution is (4.8-5.2): (1.5-2.5): (0.8-1.2): (9-12): (9-12): (5-5.2).
Preferably, the mass ratio of the bovine serum albumin, tween, polyethylene glycol, the amphotropic antibody blocker, the resin and the rabbit anti-human polyclonal antibody in the sample pad treatment solution is 5:2:1:10:10:5; and/or the sample dilution contains 0.2w/v% sodium sulphide.
Preferably, tween contained in the sample diluent and/or the sample pad treatment solution is tween 20; the polyethylene glycol contained in the sample diluent and/or the sample pad treatment liquid is polyethylene glycol 20000; the surfactant is ethylenediamine tetraacetic acid; the biological preservative is a liquid biological preservative Proclin300; and/or the resin is polyhydroxyethyl methacrylate.
Preferably, the sample diluent is prepared by using PB buffer solution with the concentration of 0.01-0.03 mol/L, pH of 7.4 as a solvent; and/or the sample pad treatment solution is prepared by taking Tris-HCl buffer solution with the concentration of 0.08-0.12 mol/L, pH of 7.4 as a solvent.
Preferably, the immunochromatographic test strip is a colloidal gold immunochromatographic test strip, and the colloidal gold immunochromatographic test strip comprises a bottom plate and a rubber strip arranged on the bottom plate; the adhesive tape comprises a sample pad, a colloidal gold marked binding pad, a reaction film coated with an antibody and a water absorption pad which are sequentially overlapped; the reaction film comprises a detection line T line and a quality control line C line.
The present invention provides in a second aspect a method for preparing an immunochromatographic test cassette according to the first aspect, characterized in that the method comprises the steps of:
(1) Preparing the sample diluent;
(2) Preparing the sample pad treatment liquid;
(3) Providing a sample pad, soaking the sample pad in the sample pad treatment liquid, and drying to obtain a sample pad treated by the sample pad treatment liquid;
(4) And assembling the sample pad treated by the sample pad treatment liquid with the combination pad, the reaction membrane, the water absorption pad and the bottom plate to form an immunochromatographic test strip, thereby obtaining the immunochromatographic test strip box comprising the immunochromatographic test strip and the sample diluent.
Preferably, soaking is carried out for 2-8 hours; the drying is carried out for 15-20 hours at 37 ℃; the sample pad is made of glass fiber; the reaction membrane is a nitrocellulose membrane; and/or the bottom plate is a polyvinyl chloride bottom plate.
The invention provides in a third aspect the use of an immunochromatographic test strip cassette according to the first aspect for eliminating non-specific interference of animal faeces samples in immunochromatographic assays.
In a fourth aspect, the present invention provides the use of an immunochromatographic test strip cassette according to the first aspect of the present invention for the detection of feline panleukopenia virus.
Compared with the prior art, the invention has at least the following beneficial effects:
(1) The components such as fecal bile pigment, urinary bile pigment, heavy metal, bacterial protein, and desquamation cell protein in animal feces components can influence the experimental detection result; the prior art cannot well solve the false positive result caused by the non-specific adsorption; the immunochromatography test paper box of the invention is matched with a sample pad treatment liquid by adopting a sample diluent with a proper formula, in particular Na therein 2 S, resin, HBR, RBC and other components are not necessary, so that interference factors in feces such as fecal bile pigment, urine bile pigment, heavy metal, bacterial protein, abscission cell protein and the like can be effectively blocked, nonspecific interference of animal feces samples in immunochromatography detection can be eliminated, the sensitivity and accuracy of detection are effectively improved, and the sample coincidence rate can be up to more than 90%.
(2) In some preferred embodiments, the sample diluent contains 0.19-0.22 w/v% sodium sulfide and the mass ratio of bovine serum albumin, tween, polyethylene glycol, an amphotropic antibody blocker, resin and rabbit anti-human polyclonal antibody contained in the sample pad treatment solution is (4.8-5.2): (1.5-2.5): (0.8-1.2): (9-12): (9-12): (5-5.2) and can be matched with each other, so that interference factors in feces such as fecal bile pigment, urine bile pigment, heavy metals, bacterial proteins and the like can be sealed to the greatest extent, nonspecific interference of animal feces samples in immunochromatography detection is eliminated more obviously, the sensitivity and accuracy of detection are improved effectively, and the sample coincidence rate can reach more than 95%.
Drawings
FIG. 1 is a flow chart of preparing an immunochromatographic test strip cassette in accordance with some embodiments of the present invention.
FIG. 2 is a graph showing false positive results of 10 negative samples and 10 immunochromatographic test strips tested in 10 immunochromatographic test cassettes of comparative example 1.
FIG. 3 is a graph showing the positive results of 10 positive samples tested in 10 immunochromatographic test cassettes of comparative example 1 and 10 immunochromatographic test strips.
FIG. 4 is a graph showing false positive results of 10 negative samples tested with 10 immunochromatographic test strips of comparative example 2.
FIG. 5 is a graph showing the positive results of 10 positive samples tested with 10 immunochromatographic test strips of comparative example 2.
FIG. 6 is a graph showing false positive results of 10 negative samples and 10 immunochromatographic test strips tested in 10 immunochromatographic test cassettes of comparative example 3.
FIG. 7 is a graph showing the positive results of 10 positive samples tested in 10 immunochromatographic test cassettes of comparative example 3 and 10 immunochromatographic test strips.
FIG. 8 is a graph showing the negative results of 10 negative samples tested with 10 immunochromatographic test cassettes in example 1 and 10 immunochromatographic test strips.
FIG. 9 is a graph showing the positive results of 10 positive samples tested in 10 immunochromatographic test cassettes in example 1 and 10 immunochromatographic test strips.
In fig. 2 to 9, the direction from right to left is the chromatographic direction.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, the technical solutions of the present invention will be clearly and completely described below in connection with the embodiments of the present invention. It will be apparent that the described embodiments are some, but not all, embodiments of the invention. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
The present invention provides in a first aspect an immunochromatographic test strip cartridge comprising an immunochromatographic test strip and a sample diluent; the sample diluent contains 0.15-0.25 w/v% (e.g., 0.15w/v%, 0.16w/v%, 0.17w/v%, 0.18w/v%, 0.19w/v%, 0.2w/v%, 0.21w/v%, 0.22w/v%, 0.23w/v%, 0.24w/v%, or 0.25 w/v%) tween, 0.05-0.15 w/v% (e.g., 0.05w/v%, 0.06w/v%, 0.07w/v%, 0.08w/v%, 0.09w/v%, 0.1w/v%, 0.11w/v%, 0.12w/v%, 0.13w/v%, 0.14w/v%, or 0.15 w/v%) polyethylene glycol (PEG), 0.05-0.15 w/v% (e.g., 0.05w/v%, 0.06w/v%, 0.07w/v%, 0.08w/v%, 0.09w/v% >, 0.09 w/v%) 0.1w/v%, 0.11w/v%, 0.12w/v%, 0.13w/v%, 0.14w/v%, or 0.15 w/v%) of a surfactant, 0.05 to 0.15w/v% (e.g., 0.05w/v%, 0.06w/v%, 0.07w/v%, 0.08w/v%, 0.09w/v%, 0.1w/v%, 0.11w/v%, 0.12w/v%, 0.13w/v%, 0.14w/v%, or 0.15 w/v%) of a biological preservative, and 0.15 to 0.25w/v% (e.g., 0.15w/v%, 0.16w/v%, 0.17w/v%, 0.18w/v%, 0.19w/v%, 0.2w/v%, 0.21w/v%, 0.22w/v%, 0.23w/v%, 0.24w/v%, or 0.25 w/v%) of sodium sulfide (Na% 2 S) S; the sample pad of the immunochromatographic test strip is treated by a sample pad treatment liquid, namely the sample pad of the immunochromatographic test strip is treated by the sample pad treatment liquid to obtain the immunochromatographic test strip; the sample pad treatment solution contains 0.45-0.55 w/v% (e.g., 0.45w/v%, 0.46w/v%, 0.47w/v%, 0.48w/v%, 0.49w/v%, 0.5w/v%, 0.51w/v%, 0.52w/v%, 0.53w/v%, 0.54w/v%, or 0.55 w/v%) Bovine Serum Albumin (BSA), 0.15-0.25 w/v% (e.g., 0.15w/v%, 0.16w/v%, 0.17w/v%, 0.18w/v%, 0.19w/v%, 0.2w/v%, 0.21w/v%, 0.22w/v%, 0.23w/v%, 0.24w/v%, or 0.25 w/v%) tween 0.05-0.15 w/v% (e.g., 0.05w/v%, 0.06w/v%, 0.07w/v%, 0.08w/v%, 0.09 w/v)1.11 w/v%, 0.12w/v%, 0.13w/v%, 0.14w/v%, or 0.15 w/v%) polyethylene glycol (PEG), 0.8-1.5 w/v% (e.g., 0.8w/v%, 0.9w/v%, 1w/v%, 1.1w/v%, 1.2w/v%, 1.3w/v%, 1.4w/v%, or 1.5 w/v%) an isophilic antibody blocker (HBR), 0.8-1.5 w/v% (e.g., 0.8w/v%, 0.9w/v%, 1w/v%, 1.1w/v%, 1.2w/v%, 1.3w/v%, 1.4w/v%, or 1.5 w/v%) resin, and 0.45-0.55 w/v% (e.g., 0.45w/v%, 0.46w/v%, 0.47w/v%, 0.48w/v%, 0.49w/v%, 0.52.52% or 0.52% of an anti-human antibody; in the present invention, the resin is a liquid resin having a boiling point of not more than 95 ℃ and being miscible with water; the sample pad treatment fluid is blocked by adopting the heterophilic antibody blocking agent (HBR), and the HBR serving as the heterophilic antibody blocking agent has the advantages that only a small amount of protein is needed for blocking, a detection signal is not weakened, and more false positives can be blocked by the HBR compared with the traditional method; HBR can block all species (such as anti-rabit, anti-coat, anti-mouse, and anti-mouse, etc.), can block all subtypes of anti-mouse monoclonal antibodies (such as anti-mouse Ig 1, anti-mouse IgG2, etc.), and RBC has great effect on eliminating animal protein interference; the sources of tween, polyethylene glycol (PEG), bovine Serum Albumin (BSA), surfactant, biological preservative, bovine Serum Albumin (BSA), amphotropic antibody blocking agent (HBR), resin, rabbit anti-human polyclonal antibody (RBC) and the like are not particularly limited, and products which can be directly purchased in the market or synthesized by the prior method can be adopted; in the present invention, "w/v%" means the unit of the concentration (final concentration) of the component contained in the sample dilution and the sample pad treatment solution, and "g/100mL".
The immunochromatography test paper box of the invention is matched with a sample pad treatment liquid by adopting a sample diluent with a proper formula, in particular Na therein 2 S, resin, HBR, RBC and other components are not necessary, and besides Na in the sample diluent 2 In addition to the great influence of S concentration on the result, the sample pad treatment solution needs to have proper concentration ratio among the components, especially between resin, HBR, RBC and bovine serum albuminThe invention has proper concentration ratio, and can effectively seal interference factors in feces such as fecal bile pigment, urine bile pigment, heavy metal, bacterial protein, abscission cell protein and the like, thereby effectively eliminating nonspecific interference of animal feces samples in immunochromatography detection, effectively improving the sensitivity and accuracy of detection; in addition, the present invention found that sodium sulfide (Na 2 S) sodium sulfide is required to be added into a sample diluent, the activity of sulfide ions can be kept in the sample diluent, resin, HBR and RBC are required to be added into the sample pad treatment liquid, and the resin, HBR and RBC are added together to have good effects.
According to some preferred embodiments, the sample pad treatment solution contains bovine serum albumin, tween, polyethylene glycol, an amphotropic antibody blocking agent, resin and rabbit anti-human polyclonal antibody in a mass ratio of (4.5-5.5): (1.5-2.5): (0.5 to 1.5): (9-12): (9-12): (4.5 to 5.5), preferably (4.8 to 5.2): (1.5-2.5): (0.8-1.2): (9-12): (9-12): (5 to 5.2), more preferably (4.8 to 5.2): (1.5-2): (1-1.2): (9-10): (9-10): (5-5.2).
According to some preferred embodiments, the sample pad treatment solution contains bovine serum albumin, tween, polyethylene glycol, an amphotropic antibody blocker, a resin and a rabbit anti-human polyclonal antibody in a mass ratio of 5:2:1:10:10:5; and/or the sample dilution contains 0.2w/v% sodium sulphide.
In some preferred embodiments, the sample diluent contains 0.19-0.22 w/v% sodium sulfide, more preferably 0.2w/v% to the mass ratio of bovine serum albumin, tween, polyethylene glycol, an amphotropic antibody blocker, a resin and a rabbit anti-human polyclonal antibody contained in the sample pad treatment solution is (4.8-5.2): (1.5-2.5): (0.8-1.2): (9-12): (9-12): (5-5.2) is more preferably matched with the mass ratio of 5:2:1:10:10:5, so that interference factors in feces such as fecal bile pigment, urinary bile pigment, heavy metal, bacterial protein and the like can be sealed to the greatest extent, nonspecific interference of animal feces samples in immunochromatography detection is eliminated more obviously, the sensitivity and accuracy of detection are improved effectively, the sample coincidence rate reaches more than 95%, and even the sample coincidence rate reaches more than 98%.
According to some preferred embodiments, the tween contained in the sample dilution and/or the sample pad treatment is tween 20; the polyethylene glycol contained in the sample diluent and/or the sample pad treatment liquid is polyethylene glycol 20000; the surfactant is ethylenediamine tetraacetic acid (EDTA); the biological preservative is a liquid biological preservative Proclin300; and/or the resin is polyhydroxyethyl methacrylate; in the invention, the liquid biological preservative Proclin300 refers to Proclin300, the sources of Proclin300 and polyhydroxyethyl methacrylate are not particularly limited, and products which can be directly purchased in the market or synthesized by the existing method can be adopted.
According to some preferred embodiments, the sample diluent is prepared by using PB buffer solution with a concentration of 0.01-0.03 mol/L (for example, 0.01, 0.02 or 0.03 mol/L) and a pH of 7.4 as a solvent; and/or the sample pad treatment solution is prepared by taking Tris-HCl buffer solution with the concentration of 0.08-0.12 mol/L (for example, 0.08, 0.09, 0.10, 0.11 or 0.12 mol/L) and the pH of 7.4 as a solvent; the PB buffer solution and the Tris-HCl buffer solution are not particularly limited, and the PB buffer solution and the Tris-HCl buffer solution can be directly purchased in the market or prepared by the existing method, so long as the concentration of the PB buffer solution is 0.01-0.03 mol/L, the pH is 7.4, and the concentration of the Tris-HCl buffer solution is 0.08-0.12 mol/L, pH and is 7.4.
In the invention, preferably, the sample diluent is prepared by taking 0.01-0.03 mol/L, pH PB buffer solution as a solvent, the sample pad treatment solution is prepared by taking 0.08-0.12 mol/L, pH Tris-HCl buffer solution as a solvent, and the use of the solvent which is more matched with each component of the sample diluent and the sample pad treatment solution can more effectively seal interference factors in excrement such as fecal bile pigment, urinary bile pigment, heavy metals, bacterial proteins, abscisic cell proteins and the like, and the sensitivity and accuracy of detection are more effectively improved; according to the invention, if the Tris-HCl buffer solution is used as a solvent for the sample diluent and the PB buffer solution is used for the sample pad treatment solution, the detection sensitivity and accuracy of the paper box can be reduced, and high false positive is caused.
According to some preferred embodiments, the immunochromatographic test strip is a colloidal gold immunochromatographic test strip, and the colloidal gold immunochromatographic test strip comprises a bottom plate and a rubber strip arranged on the bottom plate; the adhesive tape comprises a sample pad, a colloidal gold marked binding pad, a reaction film coated with an antibody and a water absorption pad which are sequentially overlapped; the reaction film comprises a detection line T line and a quality control line C line; in the invention, the sample pad can be also marked as a sample adding pad, the immunochromatographic test strip can also exist in the detection plate, namely, for example, the immunochromatographic test paper box further comprises an upper cover and a lower cover which are in sealing buckling, the upper cover is provided with a sample adding hole and a detection window, the lower cover is provided with a clamping groove, the immunochromatographic test strip is placed in the clamping groove, the position of a reaction membrane of the immunochromatographic test strip corresponds to the position of the detection window, and the position of the sample pad (sample adding pad) of the immunochromatographic test strip corresponds to the sample adding hole.
In the invention, the immunochromatography test paper box can take cat leukopenia virus as a detection object, adopts a colloidal gold-marked binding pad and a reaction membrane coated with an antibody as conventional technology in the field, and the colloidal gold-marked antibody is matched with a detection line T line coated antibody of the reaction membrane for use, so that no special requirement exists, and the immunochromatography test paper box is only suitable for cat leukopenia virus detection; in some embodiments of the present invention and comparative examples, the reaction membrane detection line T line coated antibody was cat-panleukopenia virus monoclonal antibody 7E3 (product name: FPV-7E 3) purchased from Zhengzhou Ling biosciences, inc., batch 20220416, the colloidal gold labeled antibody was cat-panleukopenia virus monoclonal antibody 5G6 (product name: FPV-5G 6), purchased from Zhengzhou Ling biosciences, inc., batch 20220221, FPV-7E3 paired with FPV-5G6, and the reaction membrane quality control line C line coated antibody was sheep anti-mouse antibody (sheep anti-mouse IGG antibody).
The present invention provides in a second aspect a method for preparing an immunochromatographic test cassette according to the first aspect, characterized in that the method comprises the steps of:
(1) Preparing the sample diluent;
(2) Preparing the sample pad treatment liquid;
(3) Providing a sample pad, soaking the sample pad in the sample pad treatment liquid, and drying (namely baking) the sample pad to obtain a sample pad treated by the sample pad treatment liquid; in the present invention, for example, the sample pad may be obtained from glass fibers, and the glass fibers are not particularly limited, and for example, a density of 0.15 to 0.65g/cm may be used 3 The glass fiber membrane is made of glass fiber as a sample pad, which is a conventional technology in the field; specifically, the glass fiber is made into a glass fiber film and cut into a proper size to obtain a whole sample pad (the whole sample pad can be cut into a plurality of sample pads for an immunochromatographic test strip), and then the whole sample pad is soaked in the sample pad treatment liquid Calculating the dosage of the sample pad treatment liquid; then, placing the soaked sample pad in a baking oven at 37 ℃, and enabling the sample pad to reach a stable state after the sample pad is placed in the baking oven for 15-20 hours, so as to obtain the sample pad treated by the sample pad treatment liquid; under the condition that the sample pad is not used, the sample pad treated by the sample pad treatment liquid can be put into a sealing bag and added with a drying agent for preservation, so that the use time of the sample pad is prolonged.
(4) Assembling the sample pad treated by the sample pad treatment liquid with the binding pad, the reaction membrane, the water absorption pad and the bottom plate to form an immunochromatography test strip; preferably, the bonding pad is a colloidal gold-labeled bonding pad; in the invention, the binding pad marked by colloidal gold is also marked as a gold mark pad, the invention carries out the colloidal gold marking on the binding pad (such as the binding pad made of glass fiber film) as a routine technology in the field, for example, colloidal gold solution (such as 100OD value colloidal gold solution), buffer solution and marked antibody are mixed uniformly and then coupled for 2 hours and then sealed for 1 hour, after centrifugal re-dissolution, the solution is sprayed on the binding pad, and then the binding pad is put into a baking oven for 4 hours and cut to obtain the binding pad marked by colloidal gold (gold mark pad); in the invention, the reaction membrane is coated with an antibody, the reaction membrane comprises a detection line T line and a quality control line C line, and the preparation of the coated antibody is a conventional technology in the field, for example, after preparing a coating solution, a film dividing instrument is used for dividing the coated C line and T line antibody, and the coated C line and T line antibody are dried for standby.
According to some preferred embodiments, the soaking is performed for a period of 2-8 hours (e.g., 2, 3, 4, 5, 6, 7, or 8 hours); preferably, soaking is carried out at room temperature (for example, at room temperature of 15-30 ℃) for 2-8 hours; the drying is carried out for 15-20 h (for example, 15, 16, 17, 18, 19 or 20 h) at 37 ℃; the sample pad is made of glass fiber; the reaction membrane is a nitrocellulose membrane (NC membrane); and/or the base plate is a polyvinyl chloride base plate (PVC base plate).
The invention provides in a third aspect the use of an immunochromatographic test strip cassette according to the first aspect for eliminating non-specific interference of animal faeces samples in immunochromatographic assays; preferably, for each immunochromatographic test paper box, 400-600 mu L of prepared sample diluent is used, the animal faeces sample is diluted in 400-600 mu L of sample diluent to obtain a detection sample, for each immunochromatographic test paper strip, the dosage of the detection sample is 55-65 mu L, more preferably, for each immunochromatographic test paper box, 500 mu L of prepared sample diluent is used, and the dosage of the corresponding detection sample is 60 mu L/strip; the invention does not limit the amount of the animal feces sample before dilution, and the animal feces sample can be taken according to the conventional operation in the field, for example, a cotton swab is used for dipping a proper amount of animal feces.
In a fourth aspect, the present invention provides the use of an immunochromatographic test strip cassette according to the first aspect of the present invention for the detection of feline panleukopenia virus.
The invention also provides a sample pad treatment liquid for treating the sample pad of the immunochromatographic test paper box, which contains 0.45-0.55 w/v% of bovine serum albumin, 0.15-0.25 w/v% of tween, 0.05-0.15 w/v% of polyethylene glycol, 0.8-1.5 w/v% of an amphotropic antibody blocker, 0.8-1.5 w/v% of resin and 0.45-0.55 w/v% of rabbit anti-human polyclonal antibody; the sample pad treatment liquid is matched with a sample diluent for use, and the sample diluent contains 0.15-0.25 w/v% of tween, 0.05-0.15 w/v% of polyethylene glycol, 0.05-0.15 w/v% of surfactant, 0.05-0.15 w/v% of biological preservative and 0.15-0.25 w/v% of sodium sulfide; preferably, the mass ratio of the bovine serum albumin, tween, polyethylene glycol, the amphotropic antibody blocking agent, the resin and the rabbit anti-human polyclonal antibody contained in the sample pad treatment solution is (4.8-5.2): (1.5-2.5): (0.8-1.2): (9-12): (9-12): (5-5.2), more preferably 5:2:1:10:10:5; preferably, the sample diluent contains 0.2w/v% sodium sulfide; preferably, tween contained in the sample diluent and/or the sample pad treatment solution is tween 20; the polyethylene glycol contained in the sample diluent and/or the sample pad treatment liquid is polyethylene glycol 20000; the surfactant is ethylenediamine tetraacetic acid; the biological preservative is a liquid biological preservative Proclin300; the resin is polyhydroxyethyl methacrylate; the sample diluent is prepared by taking PB buffer solution with the concentration of 0.01-0.03 mol/L, pH of 7.4 as a solvent; and/or the sample pad treatment solution is prepared by taking Tris-HCl buffer solution with the concentration of 0.08-0.12 mol/L, pH of 7.4 as a solvent.
The invention also provides a sample diluent for the immunochromatographic test paper box, which comprises 0.15-0.25 w/v% of tween, 0.05-0.15 w/v% of polyethylene glycol, 0.05-0.15 w/v% of surfactant, 0.05-0.15 w/v% of biological preservative and 0.15-0.25 w/v% of sodium sulfide (Na 2 S) S; the sample diluent is matched with a sample pad treatment liquid for treating a sample pad of an immunochromatographic test paper box, and the sample pad treatment liquid contains 0.45-0.55 w/v% of bovine serum albumin, 0.15-0.25 w/v% of tween, 0.05-0.15 w/v% of polyethylene glycol, 0.8-1.5 w/v% of an ampholytic antibody blocker, 0.8-1.5 w/v% of resin and 0.45-0.55 w/v% of rabbit anti-human polyclonal antibody; preferably, the mass ratio of the bovine serum albumin, tween, polyethylene glycol, the amphotropic antibody blocking agent, the resin and the rabbit anti-human polyclonal antibody contained in the sample pad treatment solution is (4.8-5.2): (1.5-2.5): (0.8-1.2): (9-12): (9-12): (5-5.2), more preferably 5:2:1:10:10:5; preferably, the sample diluent contains 0.2w/v% sodium sulfide; preferably, tween contained in the sample diluent and/or the sample pad treatment solution is tween 20; the polyethylene glycol contained in the sample diluent and/or the sample pad treatment liquid is polyethylene glycol 20000; the surfactant is ethylenediamine tetraacetic acid; the biological preservative is a liquid biological preservative Proclin300; the resin is polyhydroxyethyl methacrylate; the sample diluent is prepared by taking PB buffer solution with the concentration of 0.01-0.03 mol/L, pH of 7.4 as a solvent; and/or the sample pad treatment solution is prepared by taking Tris-HCl buffer solution with the concentration of 0.08-0.12 mol/L, pH of 7.4 as a solvent.
The invention will be further illustrated by way of example, but the scope of the invention is not limited to these examples. The present invention is capable of other and further embodiments and its several details are capable of modification and variation in accordance with the present invention, as will be apparent to those skilled in the art, without departing from the spirit and scope of the invention as defined in the appended claims.
Example 1
An immunochromatographic test paper box comprises an immunochromatographic test paper strip and a sample diluent; wherein tween 20, polyethylene glycol (PEG 20000), surfactant (EDTA), biological preservative (Proclin 300) and Na were mixed with a PB buffer at a concentration of 0.02mol/L, pH =7.4 2 S is prepared into a sample diluent which contains 0.2w/v% Tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 0.1w/v% surfactant (EDTA), 0.1w/v% biological preservative (Proclin 300) and 0.2w/v% sodium sulfide (Na) 2 S), the formulation is shown in table 1 below.
TABLE 1
The preparation of the immunochromatography test strip comprises the following steps:
(1) bovine serum albumin, tween 20, polyethylene glycol, an amphotropic antibody blocker, resin and rabbit anti-human polyclonal antibody were formulated with Tris-HCl buffer at a concentration of 0.1mol/L, pH =7.4 into sample pad treatment containing 0.5w/v% Bovine Serum Albumin (BSA), 0.2w/v% tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 1w/v% amphotropic antibody blocker (HBR), 1w/v% resin (polyhydroxyethyl methacrylate) and 0.5w/v% rabbit anti-human polyclonal antibody (RBC) as shown in table 2 below.
TABLE 2
(2) Glass fiber membrane is made of glass fiber, and then the sample pad is obtained; and soaking the sample pads in the sample pad treatment liquid for 5 hours, wherein the dosage of the sample pad treatment liquid required for soaking each sample pad is 5mL, then placing the soaked sample pads in a 37 ℃ oven, and drying (baking) the sample pads in the oven for 15 hours to enable the sample pads to reach a stable state, thereby obtaining the sample pads treated by the sample pad treatment liquid.
(3) Taking cat panleukopenia virus detection as an experimental object, manufacturing a colloidal gold-labeled binding pad and an NC film coated with an antibody; the NC film comprises a detection line T line and a quality control line C line; and assembling the sample pad treated by the sample pad treatment liquid with a colloidal gold marked binding pad, an NC film coated with an antibody, a water absorption pad and a bottom plate (PVC bottom plate) to form the immunochromatography test strip.
Example 2
Example 2 is substantially the same as example 1 except that:
bovine serum albumin, tween 20, polyethylene glycol, a blocking agent for the heterotrophic antibodies, a resin and rabbit anti-human polyclonal antibodies were formulated with Tris-HCl buffer at a concentration of 0.1mol/L, pH =7.4 into a sample pad treatment containing 0.5w/v% bovine serum albumin, 0.15w/v% tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 1w/v% blocking agent for the heterotrophic antibodies (HBR), 1w/v% resin (polyhydroxyethyl methacrylate) and 0.45w/v% rabbit anti-human polyclonal antibody (RBC).
Example 3
Example 3 is substantially the same as example 1 except that:
bovine serum albumin, tween 20, polyethylene glycol, a blocking agent for the heterotrophic antibodies, a resin and rabbit anti-human polyclonal antibodies were formulated with Tris-HCl buffer at a concentration of 0.1mol/L, pH =7.4 into a sample pad treatment containing 0.55w/v% bovine serum albumin, 0.2w/v% tween 20, 0.15w/v% polyethylene glycol (PEG 20000), 1w/v% blocking agent for the heterotrophic antibodies (HBR), 1w/v% resin (polyhydroxyethyl methacrylate) and 0.55w/v% rabbit anti-human polyclonal antibody (RBC).
Example 4
Example 4 is substantially the same as example 1 except that:
bovine serum albumin, tween 20, polyethylene glycol, a blocking agent for the heterotrophic antibodies, a resin and rabbit anti-human polyclonal antibodies were formulated with Tris-HCl buffer at a concentration of 0.1mol/L, pH =7.4 into a sample pad treatment containing 0.48w/v% bovine serum albumin, 0.15w/v% tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 1w/v% blocking agent for the heterotrophic antibodies (HBR), 0.9w/v% resin (polyhydroxyethyl methacrylate) and 0.5w/v% rabbit anti-human polyclonal antibody (RBC).
Example 5
Example 5 is substantially the same as example 1 except that:
bovine serum albumin, tween 20, polyethylene glycol, a blocking agent for the heterotrophic antibodies, a resin and rabbit anti-human polyclonal antibodies were formulated with Tris-HCl buffer at a concentration of 0.1mol/L, pH =7.4 into a sample pad treatment containing 0.52w/v% bovine serum albumin, 0.2w/v% tween 20, 0.12w/v% polyethylene glycol (PEG 20000), 0.9w/v% blocking agent for the heterotrophic antibodies (HBR), 1w/v% resin (polyhydroxyethyl methacrylate) and 0.52w/v% rabbit anti-human polyclonal antibody (RBC).
Example 6
Example 6 is substantially the same as example 1 except that:
bovine serum albumin, tween 20, polyethylene glycol, a blocking agent for the heterotrophic antibodies, a resin and rabbit anti-human polyclonal antibodies were formulated with Tris-HCl buffer at a concentration of 0.1mol/L, pH =7.4 into a sample pad treatment containing 0.3w/v% bovine serum albumin, 0.2w/v% tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 0.5w/v% blocking agent for the heterotrophic antibodies (HBR), 1w/v% resin (polyhydroxyethyl methacrylate) and 0.3w/v% rabbit anti-human polyclonal antibody (RBC).
Example 7
Example 7 is substantially the same as example 1 except that:
Bovine serum albumin, tween 20, polyethylene glycol, a blocking agent for the heterotrophic antibodies, a resin and rabbit anti-human polyclonal antibodies were formulated with Tris-HCl buffer at a concentration of 0.1mol/L, pH =7.4 into a sample pad treatment containing 0.4w/v% bovine serum albumin, 0.2w/v% tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 0.8w/v% blocking agent for the heterotrophic antibodies (HBR), 1w/v% resin (polyhydroxyethyl methacrylate) and 0.4w/v% rabbit anti-human polyclonal antibody (RBC).
Example 8
Example 8 is substantially the same as example 1 except that:
bovine serum albumin, tween 20, polyethylene glycol, a blocking agent for the heterotrophic antibodies, a resin and rabbit anti-human polyclonal antibodies were formulated with Tris-HCl buffer at a concentration of 0.1mol/L, pH =7.4 into a sample pad treatment containing 0.6w/v% bovine serum albumin, 0.2w/v% tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 1.5w/v% blocking agent for the heterotrophic antibodies (HBR), 1w/v% resin (polyhydroxyethyl methacrylate) and 0.5w/v% rabbit anti-human polyclonal antibody (RBC).
Example 9
Example 9 is substantially the same as example 1 except that:
bovine serum albumin, tween 20, polyethylene glycol, a blocking agent for the heterotrophic antibodies, a resin and a rabbit anti-human polyclonal antibody were formulated with Tris-HCl buffer at a concentration of 0.1mol/L, pH =7.4 into a sample pad treatment containing 0.35w/v% bovine serum albumin, 0.2w/v% tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 0.5w/v% blocking agent for the heterotrophic antibodies (HBR), 0.5w/v% resin (polyhydroxyethyl methacrylate) and 0.6w/v% rabbit anti-human polyclonal antibody (RBC).
Example 10
Example 10 is substantially the same as example 1 except that:
tween 20, polyethylene glycol (PEG 20000), surfactant (EDTA), biological preservative (Proclin 300) and sodium sulfide (Na) were mixed with PB buffer at a concentration of 0.02mol/L, pH =7.4 2 S) preparing a sample diluent which contains 0.2w/v% Tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 0.1w/v% surfactant (EDTA), 0.1w/v% biological preservative (Proclin 300) and 0.15w/v% sodium sulfide (Na) 2 S）。
Example 11
Example 11 is substantially the same as example 1 except that:
with a concentration of 0.02mol/L, pH =7.4 PB buffer Tween 20, polyethylene glycol (PEG 20000), surfactant (EDTA), biological preservative (Proclin 300) and sodium sulphide (Na 2 S) preparing a sample diluent which contains 0.2w/v% Tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 0.1w/v% surfactant (EDTA), 0.1w/v% biological preservative (Proclin 300) and 0.25w/v% sodium sulfide (Na) 2 S）。
Comparative example 1
Comparative example 1 is substantially the same as example 1 except that:
tween 20, polyethylene glycol (PEG 20000), surfactant (EDTA) and biological preservative (Proclin 300) were formulated with a PB buffer at a concentration of 0.02mol/L, pH =7.4 as a sample dilution containing 0.2w/v% tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 0.1w/v% surfactant (EDTA) and 0.1w/v% biological preservative (Proclin 300).
Bovine serum albumin, tween 20 and polyethylene glycol were formulated with Tris-HCl buffer at a concentration of 0.1mol/L, pH =7.4 as a sample pad treatment containing 0.5w/v% bovine serum albumin, 0.2w/v% tween 20 and 0.1w/v% polyethylene glycol (PEG 20000).
Comparative example 2
Comparative example 2 is substantially the same as example 1 except that:
bovine serum albumin, tween 20 and polyethylene glycol were formulated with Tris-HCl buffer at a concentration of 0.1mol/L, pH =7.4 as a sample pad treatment containing 0.5w/v% bovine serum albumin, 0.2w/v% tween 20 and 0.1w/v% polyethylene glycol (PEG 20000).
Comparative example 3
Comparative example 3 is substantially the same as example 1 except that:
tween 20, polyethylene glycol (PEG 20000), surfactant (EDTA) and biological preservative (Proclin 300) were formulated with a PB buffer at a concentration of 0.02mol/L, pH =7.4 as a sample dilution containing 0.2w/v% tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 0.1w/v% surfactant (EDTA) and 0.1w/v% biological preservative (Proclin 300).
Comparative example 4
Comparative example 4 is substantially the same as example 1 except that:
Tween 20, polyethylene glycol (PEG 20000), surfactant (EDTA), biological preservative (Proclin 300) and Na were buffered with Tris-HCl buffer at a concentration of 0.1mol/L, pH =7.4 2 S is prepared into a sample diluent which contains 0.2w/v% Tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 0.1w/v% surfactant (EDTA), 0.1w/v% biological preservative (Proclin 300) and 0.2w/v% sodium sulfide (Na) 2 S）。
Bovine serum albumin, tween 20, polyethylene glycol, a blocking agent for the heterotrophic antibodies, a resin and rabbit anti-human polyclonal antibodies were formulated with PB buffer at a concentration of 0.02mol/L, pH =7.4 into a sample pad treatment containing 0.5w/v% Bovine Serum Albumin (BSA), 0.2w/v% tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 1w/v% blocking agent for the heterotrophic antibodies (HBR), 1w/v% resin (polyhydroxyethyl methacrylate) and 0.5w/v% rabbit anti-human polyclonal antibody (RBC).
Comparative example 5
Comparative example 5 is substantially the same as example 1 except that:
tween 20, polyethylene glycol (PEG 20000), surfactant (EDTA), biological preservative (Proclin 300), na were mixed with PB buffer at a concentration of 0.02mol/L, pH =7.4 2 S and resin to prepare a sample diluent, wherein the sample diluent contains 0.2w/v% Tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 0.1w/v% surfactant (EDTA), 0.1w/v% biological preservative (Proclin 300), 0.2w/v% sodium sulfide (Na) 2 S) and 1w/v% resin (polyhydroxyethyl methacrylate).
Bovine serum albumin, tween 20, polyethylene glycol, a blocking agent for antibodies to isophilic antibodies and rabbit anti-human polyclonal antibodies were formulated with Tris-HCl buffer at a concentration of 0.1mol/L, pH =7.4 into a sample pad treatment containing 0.5w/v% bovine serum albumin, 0.2w/v% tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 1w/v% blocking agent for antibodies to isophilic antibodies (HBR) and 0.5w/v% rabbit anti-human polyclonal antibodies (RBC).
Comparative example 6
Comparative example 6 is substantially the same as example 1 except that:
tween 20, polyethylene glycol (PEG 20000), surfactant (EDTA) and Proclin300 were formulated with PB buffer at a concentration of 0.02mol/L, pH =7.4 to give a sample dilution containing 0.2w/v% Tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 0.1w/v% surfactant (EDTA) and 0.1w/v% biological preservative (Proclin 300).
Bovine serum albumin, tween 20, polyethylene glycol, a blocking agent for the heterotrophic antibodies, a resin, a rabbit anti-human polyclonal antibody and sodium sulfide were formulated with Tris-HCl buffer at a concentration of 0.1mol/L, pH =7.4 into a sample pad treatment containing 0.5w/v% bovine serum albumin, 0.2w/v% tween 20, 0.1w/v% polyethylene glycol (PEG 20000), 1w/v% blocking agent for the heterotrophic antibodies (HBR), 1w/v% resin (polyhydroxyethyl methacrylate), 0.5w/v% rabbit anti-human polyclonal antibody (RBC) and 0.2w/v% sodium sulfide (Na 2 S）。
Performance test 1
The detection method comprises the following steps: and (3) taking cat leukopenia virus detection as an experimental object, diluting a cat fecal sample into 500 mu L of sample diluent to obtain detection samples, wherein the use amount of the detection samples corresponding to each immunochromatography test strip is 60 mu L, and judging the result after waiting for 15 min.
10 negative samples were tested with 10 immunochromatographic test cassettes of comparative example 1, and 10 immunochromatographic test strips showed false positives, and the results are shown in FIG. 2.
10 positive samples were tested with 10 immunochromatographic test cassettes of comparative example 1, and all of the 10 immunochromatographic test strips showed positive results, as shown in FIG. 3.
10 negative samples were tested with 10 immunochromatographic test strips of comparative example 2, and the 10 immunochromatographic test strips showed false positives, and the results are shown in FIG. 4.
10 positive samples were tested by using 10 immunochromatographic test paper boxes in comparative example 2, and the results shown by 10 immunochromatographic test paper strips are positive, and the results are shown in FIG. 5.
10 negative samples were tested with 10 immunochromatographic test cassettes of comparative example 3, and 10 immunochromatographic test strips showed false positives, and the results are shown in FIG. 6.
10 positive samples were tested with 10 immunochromatographic test cassettes of comparative example 3, and 10 immunochromatographic test strips showed positive results, and the results are shown in FIG. 7.
10 negative samples were tested with 10 immunochromatographic test cassettes of example 1, and all of the 10 immunochromatographic test strips showed negative results, as shown in FIG. 8.
10 positive samples were tested with 10 immunochromatographic test cassettes of example 1, and all of the 10 immunochromatographic test strips showed positive results, as shown in FIG. 9.
Performance detection II
The detection method comprises the following steps: taking cat leukopenia virus detection as an experimental object, diluting a cat fecal sample into 500 mu L of sample diluent to obtain detection samples, judging results after waiting 15min, specifically 100 negative samples from 100 immunochromatographic test paper boxes in examples 1-11 and comparative examples 1-6 respectively, wherein the negative samples are obtained from cat individuals with PCR detection (virus nucleic acid detection) of a pet hospital, the results of false positive rates are shown in table 3, and the results of coincidence rate with the nucleic acid detection results are shown in table 3.
TABLE 3 Table 3
The invention is not described in detail in the area of the technicians in this field known technology, the technicians in this field can choose according to the needs.
Finally, it should be noted that: the above embodiments are only for illustrating the technical solution of the present invention, and are not limiting; although described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical scheme described in the foregoing embodiments can be modified or some technical features thereof can be replaced by equivalents; such modifications and substitutions do not depart from the spirit and scope of the technical solutions of the embodiments of the present invention.
Claims (10)
1. An immunochromatographic test paper cassette, which is characterized in that:
the immunochromatography test paper box comprises an immunochromatography test strip and a sample diluent;
the sample diluent comprises 0.15-0.25 w/v% of tween, 0.05-0.15 w/v% of polyethylene glycol, 0.05-0.15 w/v% of surfactant, 0.05-0.15 w/v% of biological preservative and 0.15-0.25 w/v% of sodium sulfide;
the sample pad of the immunochromatographic test strip is treated by a sample pad treatment liquid;
the sample pad treatment solution contains 0.45-0.55 w/v% of bovine serum albumin, 0.15-0.25 w/v% of tween, 0.05-0.15 w/v% of polyethylene glycol, 0.8-1.5 w/v% of an amphotropic antibody blocker, 0.8-1.5 w/v% of polyhydroxyethyl methacrylate and 0.45-0.55 w/v% of rabbit anti-human polyclonal antibody RBC.
2. The immunochromatographic test strip cassette according to claim 1, wherein:
the sample pad treatment solution contains bovine serum albumin, tween, polyethylene glycol, a heterophilic antibody blocking agent and a poly (hydroxyethyl methacrylate) and rabbit anti-human polyclonal antibody RBC in a mass ratio of (4.8-5.2): (1.5-2.5): (0.8-1.2): (9-12): (9-12): (5-5.2).
3. The immunochromatographic test strip cassette according to claim 2, wherein:
the sample pad treatment solution contains bovine serum albumin, tween, polyethylene glycol, an amphotropic antibody blocking agent, polyhydroxyethyl methacrylate and a rabbit anti-human polyclonal antibody RBC in a mass ratio of 5:2:1:10:10:5; and/or
The sample dilution contained 0.2w/v% sodium sulfide.
4. The immunochromatographic test strip cartridge according to any one of claims 1 to 3, wherein:
the tween contained in the sample diluent and/or the sample pad treatment solution is tween 20;
the polyethylene glycol contained in the sample diluent and/or the sample pad treatment liquid is polyethylene glycol 20000;
the surfactant is ethylenediamine tetraacetic acid;
the biological preservative is a liquid biological preservative Proclin300.
5. The immunochromatographic test strip cartridge according to any one of claims 1 to 3, wherein:
the sample diluent is prepared by taking PB buffer solution with the concentration of 0.01-0.03 mol/L, pH of 7.4 as a solvent; and/or
The sample pad treatment solution is prepared by taking Tris-HCl buffer solution with the concentration of 0.08-0.12 mol/L, pH of 7.4 as a solvent.
6. The immunochromatographic test strip cartridge according to any one of claims 1 to 3, wherein:
the immunochromatographic test strip is a colloidal gold immunochromatographic test strip, and comprises a bottom plate and an adhesive tape arranged on the bottom plate;
the adhesive tape comprises a sample pad, a colloidal gold marked binding pad, a reaction film coated with an antibody and a water absorption pad which are sequentially overlapped;
the reaction film comprises a detection line T line and a quality control line C line.
7. A method of making an immunochromatographic test strip cassette of claim 6, comprising the steps of:
(1) Preparing the sample diluent;
(2) Preparing the sample pad treatment liquid;
(3) Providing a sample pad, soaking the sample pad in the sample pad treatment liquid, and drying to obtain a sample pad treated by the sample pad treatment liquid;
(4) And assembling the sample pad treated by the sample pad treatment liquid with the combination pad, the reaction membrane, the water absorption pad and the bottom plate to form an immunochromatographic test strip, thereby obtaining the immunochromatographic test strip box comprising the immunochromatographic test strip and the sample diluent.
8. The method of manufacturing according to claim 7, wherein:
soaking for 2-8 hours;
the drying is carried out for 15-20 hours at 37 ℃;
the sample pad is made of glass fiber;
the reaction membrane is a nitrocellulose membrane; and/or
The bottom plate is a polyvinyl chloride bottom plate.
9. Use of the immunochromatographic test strip cassette of any one of claims 1 to 6 for eliminating non-specific interference of animal stool samples in immunochromatographic assays for non-disease diagnosis and treatment purposes.
10. Use of the immunochromatographic test cassette of any one of claims 1 to 6 for the detection of feline panleukopenia virus for non-disease diagnosis and treatment purposes.
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