CN113030076B - Vaginal secretion multi-connected detection test paper, preparation method and test method thereof - Google Patents
Vaginal secretion multi-connected detection test paper, preparation method and test method thereof Download PDFInfo
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- CN113030076B CN113030076B CN202110286602.8A CN202110286602A CN113030076B CN 113030076 B CN113030076 B CN 113030076B CN 202110286602 A CN202110286602 A CN 202110286602A CN 113030076 B CN113030076 B CN 113030076B
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- 239000012085 test solution Substances 0.000 claims description 3
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- 235000019154 vitamin C Nutrition 0.000 claims description 3
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- 208000037009 Vaginitis bacterial Diseases 0.000 description 12
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- 241000193990 Streptococcus sp. 'group B' Species 0.000 description 2
- 206010046914 Vaginal infection Diseases 0.000 description 2
- 230000002159 abnormal effect Effects 0.000 description 2
- FRTNIYVUDIHXPG-UHFFFAOYSA-N acetic acid;ethane-1,2-diamine Chemical compound CC(O)=O.CC(O)=O.CC(O)=O.CC(O)=O.NCCN FRTNIYVUDIHXPG-UHFFFAOYSA-N 0.000 description 2
- 241001148470 aerobic bacillus Species 0.000 description 2
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- 230000002074 deregulated effect Effects 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 229940032049 enterococcus faecalis Drugs 0.000 description 2
- 238000000386 microscopy Methods 0.000 description 2
- 238000011084 recovery Methods 0.000 description 2
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- 108010065152 Coagulase Proteins 0.000 description 1
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- 238000003794 Gram staining Methods 0.000 description 1
- 241000194017 Streptococcus Species 0.000 description 1
- 208000032159 Vaginal inflammation Diseases 0.000 description 1
- 201000008100 Vaginitis Diseases 0.000 description 1
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Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N21/78—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N21/78—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour
- G01N21/80—Indicating pH value
-
- 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
- G01N33/52—Use of compounds or compositions for colorimetric, spectrophotometric or fluorometric investigation, e.g. use of reagent paper and including single- and multilayer analytical elements
- G01N33/525—Multi-layer analytical elements
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- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Hematology (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- Urology & Nephrology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Plasma & Fusion (AREA)
- Biotechnology (AREA)
- Cell Biology (AREA)
- Microbiology (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention discloses a vaginal secretion multi-joint detection test paper, a preparation method and a test method thereof, wherein the vaginal secretion multi-joint detection test paper comprises dry chemical detection test paper and a color code card, and the preparation method of the vaginal secretion multi-joint detection test paper comprises the following steps: preparing a matrix strip or a test paper card; the preparation of project base paper/project reaction block also discloses a test method of the vaginal secretion multi-connected test paper, which comprises the following steps: collecting vaginal secretion; preparing a sample liquid; the sample is dripped into the dry chemical detection test paper, and the result is interpreted by the color code card, so that the reaction speed of the color developing agent or the reaction promoter in each item of the vaginal secretion multi-joint detection test paper is increased, the secondary dripping of the color developing liquid is not required, the reaction time is only 5min, the stop liquid and the color developing liquid are not required, the rapid and convenient diagnosis of an outpatient service can be met, the color code card is configured, and the visual inspection function can be realized, so that the vaginal secretion multi-joint detection test paper is used for female personal health monitoring.
Description
Technical Field
The invention relates to the technical field of in-vitro diagnostic reagents, in particular to a vaginal secretion multi-connected detection test paper, a preparation method and a test method thereof.
Background
Female lower genital tract infection is a common gynecopathy and frequently-occurring disease, the incidence frequency is high, the number of people per year exceeds 3 hundred million people, the treatment cost is high, but the cure rate is low, the recurrence rate is high, and meanwhile, the reasonable use rate of antibiotics is low, thereby promoting the vaginal microecological imbalance and seriously threatening the female life safety.
Common colpitis including Bacterial Vaginosis (BV), vulvovaginal candidiasis (VVC), trichomonas Vaginitis (TV) and aerobic colpitis (AV), laboratory diagnosis methods are mainly performed by wet film microscopy, gram staining microscopy and the like at present, but are influenced by various factors such as quality of microscopic equipment, personnel quality, experience of operators and the like, subjectivity is strong, statistics on results are difficult to judge, and in recent years, the requirements of routine detection are realized by studying functional indexes such as pH value, hydrogen peroxide (H2O 2), leucocyte Esterase (LE), sialidase (SNA), proline aminopeptidase (PIP), N-acetylglucosaminidase (NAG) and the like, and the requirements of routine detection are realized on common and multiple female genital tract infection diseases such as follows: bacterial Vaginosis (BV), vulvovaginitis (VVC) and Trichomonas Vaginitis (TV) can achieve the purposes of comprehensive, early, rapid and accurate diagnosis, complementation and even microscope inspection replacement are achieved, development is rapid, but the joint detection kit for vaginosis or the analysis test paper for vaginal secretion produced by different manufacturers on the market at present are complex in operation, different color development liquids and stop liquids need to be dripped in different time in specific projects, and the mixed liquid is easy to disturb, misoperation is caused, and the reaction needs to be incubated for 10-15min in heating equipment at 37 ℃, so that the operation time is long, and the diagnosis efficiency is affected.
Therefore, the vaginal secretion multi-connected detection test paper which is comprehensive, accurate, quick and convenient in detection and can meet the requirements of rapid and effective diagnosis of outpatient service is an urgent requirement.
Disclosure of Invention
The invention provides a vaginal secretion multi-connected detection test paper, a preparation method and a test method thereof, and aims to solve the problems that in the prior art, vaginal inflammation detection operation is complicated, misoperation is easy to occur, reaction condition time is long, and diagnosis efficiency is affected.
A vaginal secretion multiplex test paper, comprising:
The dry chemical detection test paper comprises a plastic base material and a project base paper/project reaction block, wherein the project base paper/project reaction block is at least one of pH value base paper/reaction block, hydrogen peroxide base paper/reaction block, leucocyte zymogen paper/reaction block, lactic acid base paper/reaction block, sialidase paper/reaction block, proline aminopeptidase paper/reaction block, N-acetamido zymogen paper/reaction block, beta-glucuronidase zymogen paper/reaction block, solidification zymogen paper/reaction block, oxidation zymogen paper/reaction block and amine base paper/reaction block, and the plastic base material is one of a matrix strip or a test paper card;
And the color code card is combined with the dry chemical detection test paper.
The leucocyte zymogen paper/reaction block comprises 0.5 g/L-1.0 g/L of a first reaction substrate, 0.1 g/L-1.0 g/L of a diazonium salt first color reagent, 1.0 g/L-10 g/L of a first reaction promoter, 0.5 g/L-1.0 g/L of a first stabilizer and a first PBS buffer solution, wherein the first reaction substrate is at least one of indoxyl esters and derivatives thereof, pyrrole esters and pyrrolyl amino acid ester derivatives thereof, the first stabilizer is at least one of beta-cyclodextrin, mannitol, ethylene glycol, glycerol, ethylene diamine tetraacetic acid sodium salt and ethylene diamine tetraacetic acid potassium salt, and the first reaction promoter is at least one of sodium chloride and potassium thiocyanate;
the hydrogen peroxide base paper/reaction block comprises 0.5 g/L-1.0 g/L of peroxidase or horseradish peroxidase, 0.1 g/L-0.6 g/L of second color development agent, 0.1 g/L-1.0 g/L of second reaction accelerator, and a second buffer solution as a solvent, wherein the second color development agent is as follows: at least one of 4-aminoantipyrine, DHBS and 1, 5-dimethyl-2-phenyl-4-amino-3-pyrazolone (4-AAP), wherein the second reaction promoter is at least one of N-ethyl-N- (2-hydroxy-3-sulfopropyl) m-toluidine and 3, 5-dichloro-2-hydroxybenzenesulphonic acid, and the second buffer solution is any one of PBS buffer, citric acid-trisodium citrate buffer and boric acid-borax buffer;
the pH value base paper/reaction block comprises 0.1-0.6 g/L bromocresol green, 0.1-2 g/L surfactant and 0.1-2 g/L second stabilizer, wherein the concentration of ethanol in the solution is 10-100%, and the second stabilizer is at least one of sodium alginate, trehalose, sodium carboxymethylcellulose, polyvinylpyrrolidone and gelatin;
The beta-glucuronide zymogen paper/reaction block comprises a second reaction substrate composed of at least one of 0.1 g/L-4.0 g/L of 5-bromo-4-chloro-3-indolyl-beta-D-glucuronide sodium salt, 4-nitrophenyl-beta-D-glucopyranoside, 5-bromo-4-chloro-3-indolyl-beta-D-glucopyranoside and phenolphthalein glucuronide, 0.1 g/L-4.0 g/L of diazonium salt first color reagent, 0.2 g/L-6.0 g/L of third reaction promoter, 1 g/L-10 g/L of third stabilizing agent, the solvent is a third buffer solution, the third buffer solution is any one of PBS buffer solution, citric acid-trisodium citrate buffer solution, citric acid-TRIS buffer solution, boric acid-borax buffer solution and sodium acetate buffer solution, the third reaction promoter is at least one of magnesium chloride, zinc chloride, calcium chloride, ethylene glycol, sodium alginate, sodium salt of at least one of diamine, ethylene acetate, sodium salt of ethylene diamine and ethylene acetate, and at least one of stabilizing agents;
The proclotting paper/reaction block comprises glycyl-arginyl-4-methoxy-beta-naphthylamine, diazonium salt type first color developing agent and fourth stabilizer which are mixed solution of trehalose and sucrose, wherein the concentration of the glycyl-arginyl-4-methoxy-beta-naphthylamine is 1g/L to 10g/L, and the solvent is hydrochloric acid-TRIS buffer solution;
The N-acetylglucosaminidase paper/reaction block comprises a fourth reaction substrate composed of at least one of 0.1g/L-4g/L of 4-nitrophenyl-2-acetamido-2-deoxy-beta-D-glucopyranoside, 5-bromo-4-chloro-3-indolyl-beta-D-glucuronide, p-nitrophenol-N-acetyl-beta-D-aminoglycoside, 2-chloro-4-nitrophenyl-N-acetyl-beta-D-aminoglycoside, 4-nitrophenyl-N-acetyl-beta-D-aminoglycoside and 5- [4- (3-methoxy-benzalkene-rhodanine) ] -3-ammonium acetate-N-acetamido-beta-D-glucoside, the solvent is PBS buffer solution, the fourth reaction promoter is at least one of sodium tungstate and bovine serum albumin, and the fifth stabilizer is at least one of vitamin C, ascorbyl oxidase, glutathione and potassium iodide;
The zymolyside paper/reaction block comprises a chromogenic substrate layer and a chromogenic promoting layer, wherein the chromogenic substrate layer comprises 0.5 g/L-4.0 g/L of at least one of 5-bromo-4-chloro-3-indole-alpha-D-N-acetylneuraminic acid sodium salt, cresol blue-alpha-N-acetylneuraminic acid glycoside sodium salt, nitrobenzene-N-acetyl-alpha-sialidamide, naphthol-N-acetyl-alpha-sialidamide or 5-bromo-4 chloro-3-indolylacetylceramic acid salt, the solvent is a third buffer solution, the chromogenic substrate layer comprises 0.5 g/L-1.0 g/L of diazonium salt, the fifth reaction promoting agent is at least one of sodium tungstate, polyvinylpyrrolidone and thiols, and the sixth stabilizing agent is at least one of beta-cyclodextrin, mannitol, ethylene glycol, glycerol, ethylenediamine tetraacetic acid sodium salt, ethylenediamine tetraacetic acid salt and serum albumin;
The proline aminopeptidase paper/reaction block comprises a sixth reaction substrate prepared from at least one of L-proline-paranitroaniline trifluoroacetate, Z-glycylproline-paranitroaniline, L-proline beta-naphthamide hydrochloride and L-prolyl-paranitroaniline with the concentration of 0.4-5 g/L, and the solvent is a third buffer solution; the sixth reaction promoter is a diazonium salt reagent, and the seventh stabilizer is at least one of tartaric acid, oxalic acid tetraacetic acid sodium salt, potassium salt and maleic acid;
the lactic acid base paper/reaction block comprises 0.1-1.0 g/L peroxidase, 0.1-1.0 g/L lactic acid oxidase, a surfactant, a third color developing agent which is 3,3', 5' -tetramethyl benzidine and a solvent which is a first PBS buffer solution;
The oxidation zymogen paper/reaction block comprises a seventh reaction substrate of 1 g/L-10 g/L tetramethyl p-phenylenediamine, 0.5 g/L-5 g/L surfactant and a second buffer solution as a solvent;
the amine base paper/reaction block consists of a reaction layer and a color development layer, wherein the reaction layer is a potassium hydroxide solution with the mass fraction of 0.1% -5%; the color-developing layer contains 20 mg/L-500 mg/L bromocresol green, 0.8 g/L-2 g/L surfactant, and the concentration of ethanol in the solution is 10% -100%.
Wherein the surfactant is at least one of OP emulsifier, triton X-100, tween 20, tween 40, tween 60, tween 80 and polyethylene glycol.
Wherein the organic acid is at least one of sodium benzoate, benzoic acid, citric acid and tartaric acid with the concentration of 0.1 g/L-10 g/L.
The preparation method of the vaginal secretion multi-connected detection test paper is adopted, and the preparation method of the dry chemical detection test paper is as follows:
preparation of matrix strips or test paper cards:
Adhering the project base paper on a sheet or a coiled material made of at least one material of PET, PVC, PP, PS and ABS to form a matrix strip; the test paper card is made of at least one material of ABS, PP, modified PP and PMMA, a plurality of round holes with the diameter of 3 mm-7 mm are formed in the test paper card, the number of the round holes is consistent with that of the project reaction blocks, and each round hole is filled with the corresponding project reaction block;
Preparation of project base paper/project reaction block:
preparing a leucocyte zymogen paper/reaction block, dissolving a first reaction substrate in a first PBS buffer solution with the pH value of 6-9 and the concentration of 0.1mM-2mM, adding 1g/L-10g/L of a first stabilizer, forming a solution A 1 by 0.5g/L-5g/L of a first reaction promoter, dissolving a diazonium salt type first color reagent in an organic acid solvent containing 0.5g/L-10g/L and adding 0.1g/L-5g/L of a surfactant to form a solution B 1, mixing the solution A 1 and the solution B 1 according to the volume ratio of 1:1-9:1 to form a leucocyte esterase solution, soaking filter paper in the leucocyte esterase solution, and drying to prepare the leucocyte esterase paper block, or dripping 5-30ul of the leucocyte solution into a reaction hole of the punched filter paper, and drying to form a leucocyte esterase reaction hole;
Preparing hydrogen peroxide base paper/reaction block, dissolving peroxidase or horseradish peroxidase in a second buffer solution with a pH value of 5-8 and a concentration of 0.1mM-2mM, adding 0.1 g/L-0.6 g/L of a second color reagent and 0.1 g/L-1.0 g/L of a second reaction accelerator to prepare a hydrogen peroxide test solution, soaking filter paper in the hydrogen peroxide solution, and drying, or dripping 5-30ul of the hydrogen peroxide solution into reaction holes of the punched filter paper, and drying to form hydrogen peroxide reaction holes;
preparing pH value base paper/reaction block, dissolving bromocresol green and a surfactant in a 10% -100% ethanol solvent to form A 2 liquid, dissolving a second stabilizer in purified water or ethanol to form B 2 liquid, mixing the A 2 liquid and the B 2 liquid according to any volume ratio of 9:1-1:9 to form pH value solution, soaking filter paper in the pH value solution, and drying to obtain pH value paper block, or dripping 5-30ul of pH value solution into a reaction hole of punched filter paper, and drying to form a pH value reaction hole;
Preparing beta-glucuronidase paper/reaction block, dissolving a second reaction substrate in a third buffer solution with the pH value of 4-9 and the concentration of 0.5-10 mM, adding 1g/L-10g/L of a third stabilizer and 0.5g/L-5g/L of a third reaction accelerator to form A 3 liquid, dissolving diazonium salt type first color reagent in an organic acid solution with the concentration of 0.5g/L-10g/L to form B 3 liquid, mixing the A 3 liquid and the B 3 liquid according to the volume ratio of 1:1 to form beta-glucuronidase solution, soaking filter paper in the beta-glucuronidase solution, and drying to prepare the beta-glucuronidase paper block, or dripping 5-30ul of the beta-glucuronidase solution into a reaction hole of the punched filter paper, and drying to form the beta-glucuronidase reaction hole;
Preparing a clotting zymogen paper/reaction block, namely dissolving a third reaction substrate in a hydrochloric acid-TRIS buffer solution with the pH value of 3-7 and the concentration of 0.1-5 mM, buffering the solution, adding a fourth stabilizer, dissolving the solution to form A 4 solution, dissolving diazonium salt type first color reagent in an organic acid solution with the concentration of 0.5-10 g/L to form B 4 solution, soaking and drying filter paper in a clotting enzyme A 4 solution, soaking the filter paper in a clotting enzyme B 4 solution, and drying to obtain a clotting enzyme paper block, or dripping 5-30ul of clotting enzyme A 4 solution into a reaction hole of the punched filter paper, drying, dripping 5-30ul of clotting enzyme A 4 solution, and drying to obtain a clotting enzyme reaction hole;
Preparing N-acetylglucosaminidase paper/reaction block, dissolving 0.1g/L-4g/L of fourth reaction substrate in PBS buffer solution with pH value of 7-10 and concentration of 0.1mM-10mM, adding 1g/L-10g/L of fifth stabilizer and 0.5g/L-5g/L of fourth reaction promoter to form N-acetylglucosaminidase solution, soaking filter paper in the N-acetylglucosaminidase solution, and drying to obtain N-acetylglucosaminidase paper block, or dripping 5-30ul of N-acetylglucosaminidase solution into a reaction hole of the punched filter paper, and drying to form N-acetylglucosaminidase reaction hole;
Preparing a sialidase paper/reaction block, namely dissolving 0.5g/L-4.0g/L of a fifth reaction substrate in a buffer solution with the pH value of 3-7 and the concentration of 0.1mM-10mM, adding 1g/L-10g/L of a sixth stabilizer, dissolving 0.1g/L-10g/L of a fifth reaction promoter into a solution A 5, dissolving diazonium salt type first color reagent in an organic acid solution containing 0.1g/L-10g/L to form a solution B 5, mixing the solution A 5 and the solution B 5 according to the volume ratio of 1:1 to form a sialidase solution, soaking filter paper in the sialidase solution, and drying to prepare the sialidase solution or dripping 5-30ul of the sialidase solution into a reaction hole of the punched filter paper, and drying to form a sialidase reaction hole;
Preparing proline aminopeptidase paper/reaction block, namely dissolving 0.4g/L-5g/L of sixth reaction substrate in a buffer solution with the pH value of 6-8 and the concentration of 0.1mM-10mM, adding 1g/L-10g/L of seventh stabilizer and 0.5g/L-5g/L of sixth reaction accelerator to form proline aminopeptidase solution, soaking filter paper in the proline aminopeptidase solution, and drying to obtain the proline aminopeptidase paper block, or dripping 5-30ul of proline aminopeptidase solution into reaction holes of the punched filter paper, and drying to form proline aminopeptidase reaction holes;
Preparing lactic acid base paper/reaction block, dissolving 0.1-1.0 g/L peroxidase, 0.1-1.0 g/L lactic acid oxidase, surfactant and 3,3', 5' -tetramethyl benzidine as a third color reagent in a first PBS buffer solution with pH value of 5-7 to form lactic acid solution, soaking filter paper in the lactic acid solution and drying to obtain lactic acid paper block, or dripping 5-30ul lactic acid solution into reaction holes of the punched filter paper and drying to form lactic acid reaction holes;
Preparing oxidized zymogen paper/reaction block, dissolving 1 g/L-10 g/L of seventh reaction substrate in a second buffer solution, adding 0.5 g/L-5 g/L of surfactant to form oxidase reagent solution, soaking filter paper in the oxidase solution, and drying to obtain the oxidized zymogen paper/reaction block; or dripping 5-30ul oxidase solution into the reaction holes of the punched filter paper, and drying to form oxidase reaction holes;
Preparing amine base paper/reaction block, soaking and drying 0.1% -5% potassium hydroxide solution and filter paper to form reaction layer base paper; dissolving bromocresol green and a surfactant in a 10% -100% ethanol solvent to form A 6 liquid; dissolving a second stabilizer in purified water or ethanol to form a B 6 solution; mixing the solution A 6 and the solution B 6 according to any proportion of 9:1-0:10 to form a color developing layer solution, soaking filter paper in the color developing layer solution, drying to form color developing layer base paper, attaching the reaction layer base paper to the color developing layer base paper to form amine base paper, or dripping 5-30ul of the color developing layer solution into holes of the punched filter paper, drying to form amine test paper color developing holes, and cutting the reaction layer base paper into proper sizes and attaching the color developing holes to form amine reaction holes.
The test method adopting the vaginal secretion multi-connected test paper comprises the following steps of:
rotating the aseptic cotton swab or the aseptic swab at a fornix behind the vagina for 10-20 seconds until secretion is clearly observed to be attached to the cotton swab or the aseptic swab;
collecting samples by using sterile swabs or sterile swabs, putting the samples into soft test tubes, adding physiological saline into each sample, repeatedly rinsing, and fully eluting the samples to obtain sample liquid;
Taking out the dry chemical detection test paper, and dripping 20-30ul of samples into each project test paper hole, wherein about 1 drop of samples is obtained;
the test strip is placed on a specific instrument for detection, or the test strip is incubated for 5 minutes at 37+/-1 ℃ and then the result is interpreted by comparing with a color code card within 1 minute.
The application of the vaginal secretion multi-joint detection test paper in detecting at least one of PH value, beneficial bacteria, leucocyte, lactobacillus, BV pathogenic bacteria, gonococcus, gardnerella and campylobacter, candida albicans and trichomonas, escherichia coli and group B streptococcus, staphylococcus aureus and enterococcus faecalis and leucorrhea in vaginal secretion.
The beneficial effects of the invention are as follows: the color reagent or the reaction promoter in each item of the vaginal secretion multi-joint detection test paper provided by the invention accelerates the reaction speed, simultaneously, the secondary dropwise addition of the color developing liquid is not required, the reaction time is only 5 minutes, the stop liquid and the color developing liquid are not required, the rapid and convenient detection can be realized, the rapid and effective diagnosis requirement of an outpatient can be met, and the color code card is configured, so that the visual detection function can be realized, and the rapid and convenient detection device is also used for female personal health monitoring.
Drawings
In order to more clearly illustrate the embodiments of the invention or the technical solutions in the prior art, the drawings that are required in the embodiments or the description of the prior art will be briefly described, it being obvious that the drawings in the following description are only some embodiments of the invention, and that other drawings may be obtained according to these drawings without inventive effort for a person skilled in the art.
FIG. 1 is a schematic diagram of the dry test strip of the vaginal secretion multiplex test strip of the present invention.
FIG. 2 is a schematic diagram of the structure of a color code card of the vaginal secretion multi-test paper of the invention.
FIG. 3 is a schematic diagram showing the steps of a method for preparing a multi-test strip for vaginal secretion.
10-Dry chemical detection test paper, 20-color code card, 11-project base paper/project reaction block and 12-plastic substrate.
Detailed Description
Embodiments of the present invention are described in detail below, examples of which are illustrated in the accompanying drawings, wherein like or similar reference numerals refer to like or similar elements or elements having like or similar functions throughout. The embodiments described below by referring to the drawings are illustrative and intended to explain the present invention and should not be construed as limiting the invention.
In the description of the present invention, it should be understood that the terms "length," "width," "upper," "lower," "front," "rear," "left," "right," "vertical," "horizontal," "top," "bottom," "inner," "outer," and the like indicate orientations or positional relationships based on the orientation or positional relationships shown in the drawings, merely to facilitate describing the present invention and simplify the description, and do not indicate or imply that the devices or elements referred to must have a specific orientation, be configured and operated in a specific orientation, and therefore should not be construed as limiting the present invention. Furthermore, in the description of the present invention, the meaning of "a plurality" is two or more, unless explicitly defined otherwise.
Referring to fig. 1 to 3, the present invention provides a technical solution:
A vaginal secretion multiplex test paper, comprising:
The dry chemical test paper 10 is composed of a plastic base material 12 and a project base paper/project reaction block 11, wherein the project base paper/project reaction block 11 is at least one of a pH base paper/reaction block, a hydrogen peroxide base paper/reaction block, a leucocyte zymogen paper/reaction block, a lactic acid base paper/reaction block, a sialulopsinogen paper/reaction block, a proline aminopeptidase paper/reaction block, an N-acetaminogen zymogen paper/reaction block, a beta-glucuronidase zymogen paper/reaction block, a clotting zymogen paper/reaction block, an oxidation zymogen paper/reaction block and an amine base paper/reaction block, and the plastic base material 12 is one of a base strip or a test paper card;
a color code card 20, wherein the color code card 20 is used together with the dry chemical detection test paper 10.
Further, the leucocyte zymogen paper/reaction block comprises 0.5 g/L-1.0 g/L of a first reaction substrate, 0.1 g/L-1.0 g/L of a diazonium salt first color reagent, 1.0 g/L-10 g/L of a first reaction promoter, 0.5 g/L-1.0 g/L of a first stabilizer and a first PBS buffer solution, wherein the first reaction substrate is at least one of indoxyl esters and derivatives thereof, pyrrole esters and pyrrolyl amino acid ester derivatives thereof, the first stabilizer is at least one of beta-cyclodextrin, mannitol, ethylene glycol, glycerol, sodium ethylenediamine tetraacetate and potassium ethylenediamine tetraacetate, and the first reaction promoter is at least one of sodium chloride and potassium thiocyanate;
the hydrogen peroxide base paper/reaction block comprises 0.5 g/L-1.0 g/L of peroxidase or horseradish peroxidase, 0.1 g/L-0.6 g/L of second color development agent, 0.1 g/L-1.0 g/L of second reaction accelerator, and a second buffer solution as a solvent, wherein the second color development agent is as follows: at least one of 4-aminoantipyrine, DHBS and 1, 5-dimethyl-2-phenyl-4-amino-3-pyrazolone (4-AAP), wherein the second reaction promoter is at least one of N-ethyl-N- (2-hydroxy-3-sulfopropyl) m-toluidine and 3, 5-dichloro-2-hydroxybenzenesulphonic acid, and the second buffer solution is any one of PBS buffer, citric acid-trisodium citrate buffer and boric acid-borax buffer;
the pH value base paper/reaction block comprises 0.1-0.6 g/L bromocresol green, 0.1-2 g/L surfactant and 0.1-2 g/L second stabilizer, wherein the concentration of ethanol in the solution is 10-100%, and the second stabilizer is at least one of sodium alginate, trehalose, sodium carboxymethylcellulose, polyvinylpyrrolidone and gelatin;
The beta-glucuronide zymogen paper/reaction block comprises a second reaction substrate composed of at least one of 0.1 g/L-4.0 g/L of 5-bromo-4-chloro-3-indolyl-beta-D-glucuronide sodium salt, 4-nitrophenyl-beta-D-glucopyranoside, 5-bromo-4-chloro-3-indolyl-beta-D-glucopyranoside and phenolphthalein glucuronide, 0.1 g/L-4.0 g/L of diazonium salt first color reagent, 0.2 g/L-6.0 g/L of third reaction promoter, 1 g/L-10 g/L of third stabilizing agent, the solvent is a third buffer solution, the third buffer solution is any one of PBS buffer solution, citric acid-trisodium citrate buffer solution, citric acid-TRIS buffer solution, boric acid-borax buffer solution and sodium acetate buffer solution, the third reaction promoter is at least one of magnesium chloride, zinc chloride, calcium chloride, ethylene glycol, sodium alginate, sodium salt of at least one of diamine, ethylene acetate, sodium salt of ethylene diamine and ethylene acetate, and at least one of stabilizing agents;
The proclotting paper/reaction block comprises glycyl-arginyl-4-methoxy-beta-naphthylamine, diazonium salt type first color developing agent and fourth stabilizer which are mixed solution of trehalose and sucrose, wherein the concentration of the glycyl-arginyl-4-methoxy-beta-naphthylamine is 1g/L to 10g/L, and the solvent is hydrochloric acid-TRIS buffer solution;
The N-acetylglucosaminidase paper/reaction block comprises a fourth reaction substrate composed of at least one of 0.1g/L-4g/L of 4-nitrophenyl-2-acetamido-2-deoxy-beta-D-glucopyranoside, 5-bromo-4-chloro-3-indolyl-beta-D-glucuronide, p-nitrophenol-N-acetyl-beta-D-aminoglycoside, 2-chloro-4-nitrophenyl-N-acetyl-beta-D-aminoglycoside, 4-nitrophenyl-N-acetyl-beta-D-aminoglycoside and 5- [4- (3-methoxy-benzalkene-rhodanine) ] -3-ammonium acetate-N-acetamido-beta-D-glucoside, the solvent is PBS buffer solution, the fourth reaction promoter is at least one of sodium tungstate and bovine serum albumin, and the fifth stabilizer is at least one of vitamin C, ascorbyl oxidase, glutathione and potassium iodide;
The zymolyside paper/reaction block comprises a chromogenic substrate layer and a chromogenic promoting layer, wherein the chromogenic substrate layer comprises 0.5 g/L-4.0 g/L of at least one of 5-bromo-4-chloro-3-indole-alpha-D-N-acetylneuraminic acid sodium salt, cresol blue-alpha-N-acetylneuraminic acid glycoside sodium salt, nitrobenzene-N-acetyl-alpha-sialidamide, naphthol-N-acetyl-alpha-sialidamide or 5-bromo-4 chloro-3-indolylacetylceramic acid salt, the solvent is a third buffer solution, the chromogenic substrate layer comprises 0.5 g/L-1.0 g/L of diazonium salt, the fifth reaction promoting agent is at least one of sodium tungstate, polyvinylpyrrolidone and thiols, and the sixth stabilizing agent is at least one of beta-cyclodextrin, mannitol, ethylene glycol, glycerol, ethylenediamine tetraacetic acid sodium salt, ethylenediamine tetraacetic acid salt and serum albumin;
The proline aminopeptidase paper/reaction block comprises a sixth reaction substrate prepared from at least one of L-proline-paranitroaniline trifluoroacetate, Z-glycylproline-paranitroaniline, L-proline beta-naphthamide hydrochloride and L-prolyl-paranitroaniline with the concentration of 0.4-5 g/L, and the solvent is a third buffer solution; the sixth reaction promoter is a diazonium salt reagent, and the seventh stabilizer is at least one of tartaric acid, oxalic acid tetraacetic acid sodium salt, potassium salt and maleic acid;
the lactic acid base paper/reaction block comprises 0.1-1.0 g/L peroxidase, 0.1-1.0 g/L lactic acid oxidase, a surfactant, a third color developing agent which is 3,3', 5' -tetramethyl benzidine and a solvent which is a first PBS buffer solution;
The oxidation zymogen paper/reaction block comprises a seventh reaction substrate of 1 g/L-10 g/L tetramethyl p-phenylenediamine, 0.5 g/L-5 g/L surfactant and a second buffer solution as a solvent;
the amine base paper/reaction block consists of a reaction layer and a color development layer, wherein the reaction layer is a potassium hydroxide solution with the mass fraction of 0.1% -5%; the color-developing layer contains 20 mg/L-500 mg/L bromocresol green, 0.8 g/L-2 g/L surfactant, and the concentration of ethanol in the solution is 10% -100%.
Wherein the surfactant is at least one of OP emulsifier, triton X-100, tween 20, tween 40, tween 60, tween 80 and polyethylene glycol.
Further, the organic acid is at least one of sodium benzoate, benzoic acid, citric acid and tartaric acid with the concentration of 0.1g/L to 10 g/L.
Further, the dry chemical test paper 10 includes a card type test paper and a strip type test paper, each prepared base paper is divided into test paper strips with certain specification according to the requirement, and the test paper strips are adhered to the base paper strips, and then the test paper strips are cut to prepare the strip type test paper; and cutting each item base paper according to the round hole size of the test paper card, and punching the cut item base paper into a corresponding round hole and preparing the card type test paper by referring to the corresponding item reaction block.
Furthermore, the dry chemical test paper 10 can be visually inspected and also can be matched with a special analyzer for vaginal secretion, and the special analyzer for vaginal secretion is not described in detail in the present application because it belongs to the prior art.
The preparation method of the vaginal secretion multi-joint detection test paper is adopted, and the preparation method of the dry chemical detection test paper 10 is as follows:
S101: preparation of matrix strips or test paper cards:
adhering the project base paper on a sheet or a coiled material made of at least one material of PET, PVC, PP, PS and ABS to form a matrix strip; the test paper card is made of at least one material selected from ABS, PP, modified PP and PMMA, and is provided with a plurality of test paper cards The number of the round holes is consistent with that of the project reaction blocks, and each round hole is filled with the corresponding project reaction block;
s101: preparation of project base paper/project reaction block 11:
preparing a leucocyte zymogen paper/reaction block, dissolving a first reaction substrate in a first PBS buffer solution with the pH value of 6-9 and the concentration of 0.1mM-2mM, adding 1g/L-10g/L of a first stabilizer, forming a solution A 1 by 0.5g/L-5g/L of a first reaction promoter, dissolving a diazonium salt type first color reagent in an organic acid solvent containing 0.5g/L-10g/L and adding 0.1g/L-5g/L of a surfactant to form a solution B 1, mixing the solution A 1 and the solution B 1 according to the volume ratio of 1:1-9:1 to form a leucocyte esterase solution, soaking filter paper in the leucocyte esterase solution, and drying to prepare the leucocyte esterase paper block, or dripping 5-30ul of the leucocyte solution into a reaction hole of the punched filter paper, and drying to form a leucocyte esterase reaction hole;
Preparing hydrogen peroxide base paper/reaction block, dissolving peroxidase or horseradish peroxidase in a second buffer solution with a pH value of 5-8 and a concentration of 0.1mM-2mM, adding 0.1 g/L-0.6 g/L of a second color reagent and 0.1 g/L-1.0 g/L of a second reaction accelerator to prepare a hydrogen peroxide test solution, soaking filter paper in the hydrogen peroxide solution, and drying, or dripping 5-30ul of the hydrogen peroxide solution into reaction holes of the punched filter paper, and drying to form hydrogen peroxide reaction holes;
preparing pH value base paper/reaction block, dissolving bromocresol green and a surfactant in a 10% -100% ethanol solvent to form A 2 liquid, dissolving a second stabilizer in purified water or ethanol to form B 2 liquid, and mixing the A 2 liquid and the B 2 liquid according to a ratio of 9:1-1: 9, mixing according to any volume ratio to form a pH value solution, soaking the filter paper in the pH value solution, and drying to obtain a pH value paper block, or dripping 5-30ul of the pH value solution into the reaction holes of the punched filter paper, and drying to form pH value reaction holes;
Preparing beta-glucuronidase paper/reaction block, dissolving a second reaction substrate in a third buffer solution with the pH value of 4-9 and the concentration of 0.5-10 mM, adding 1g/L-10g/L of a third stabilizer and 0.5g/L-5g/L of a third reaction accelerator to form A 3 liquid, dissolving diazonium salt type first color reagent in an organic acid solution with the concentration of 0.5g/L-10g/L to form B 3 liquid, mixing the A 3 liquid and the B 3 liquid according to the volume ratio of 1:1 to form beta-glucuronidase solution, soaking filter paper in the beta-glucuronidase solution, and drying to prepare the beta-glucuronidase paper block, or dripping 5-30ul of the beta-glucuronidase solution into a reaction hole of the punched filter paper, and drying to form the beta-glucuronidase reaction hole;
Preparing a clotting zymogen paper/reaction block, namely dissolving a third reaction substrate in a hydrochloric acid-TRIS buffer solution with the pH value of 3-7 and the concentration of 0.1-5 mM, buffering the solution, adding a fourth stabilizer, dissolving the solution to form A 4 solution, dissolving diazonium salt type first color reagent in an organic acid solution with the concentration of 0.5-10 g/L to form B 4 solution, soaking and drying filter paper in a clotting enzyme A 4 solution, soaking the filter paper in a clotting enzyme B 4 solution, and drying to obtain a clotting enzyme paper block, or dripping 5-30ul of clotting enzyme A 4 solution into a reaction hole of the punched filter paper, drying, dripping 5-30ul of clotting enzyme A 4 solution, and drying to obtain a clotting enzyme reaction hole;
Preparing N-acetylglucosaminidase paper/reaction block, dissolving 0.1g/L-4g/L of fourth reaction substrate in PBS buffer solution with pH value of 7-10 and concentration of 0.1mM-10mM, adding 1g/L-10g/L of fifth stabilizer and 0.5g/L-5g/L of fourth reaction promoter to form N-acetylglucosaminidase solution, soaking filter paper in the N-acetylglucosaminidase solution, and drying to obtain N-acetylglucosaminidase paper block, or dripping 5-30ul of N-acetylglucosaminidase solution into a reaction hole of the punched filter paper, and drying to form N-acetylglucosaminidase reaction hole;
Preparing a sialidase paper/reaction block, namely dissolving 0.5g/L-4.0g/L of a fifth reaction substrate in a buffer solution with the pH value of 3-7 and the concentration of 0.1mM-10mM, adding 1g/L-10g/L of a sixth stabilizer, dissolving 0.1g/L-10g/L of a fifth reaction promoter into a solution A 5, dissolving diazonium salt type first color reagent in an organic acid solution containing 0.1g/L-10g/L to form a solution B 5, mixing the solution A 5 and the solution B 5 according to the volume ratio of 1:1 to form a sialidase solution, soaking filter paper in the sialidase solution, and drying to prepare the sialidase solution or dripping 5-30ul of the sialidase solution into a reaction hole of the punched filter paper, and drying to form a sialidase reaction hole;
Preparing proline aminopeptidase paper/reaction block, namely dissolving 0.4g/L-5g/L of sixth reaction substrate in a buffer solution with the pH value of 6-8 and the concentration of 0.1mM-10mM, adding 1g/L-10g/L of seventh stabilizer and 0.5g/L-5g/L of sixth reaction accelerator to form proline aminopeptidase solution, soaking filter paper in the proline aminopeptidase solution, and drying to obtain the proline aminopeptidase paper block, or dripping 5-30ul of proline aminopeptidase solution into reaction holes of the punched filter paper, and drying to form proline aminopeptidase reaction holes;
Preparing lactic acid base paper/reaction block, dissolving 0.1-1.0 g/L peroxidase, 0.1-1.0 g/L lactic acid oxidase, surfactant and 3,3', 5' -tetramethyl benzidine as a third color reagent in a first PBS buffer solution with pH value of 5-7 to form lactic acid solution, soaking filter paper in the lactic acid solution and drying to obtain lactic acid paper block, or dripping 5-30ul lactic acid solution into reaction holes of the punched filter paper and drying to form lactic acid reaction holes;
Preparing oxidized zymogen paper/reaction block, dissolving 1 g/L-10 g/L of seventh reaction substrate in a second buffer solution, adding 0.5 g/L-5 g/L of surfactant to form oxidase reagent solution, soaking filter paper in the oxidase solution, and drying to obtain the oxidized zymogen paper/reaction block; or dripping 5-30ul oxidase solution into the reaction holes of the punched filter paper, and drying to form oxidase reaction holes;
Preparing amine base paper/reaction block, soaking and drying 0.1% -5% potassium hydroxide solution and filter paper to form reaction layer base paper; dissolving bromocresol green and a surfactant in a 10% -100% ethanol solvent to form A 6 liquid; dissolving a second stabilizer in purified water or ethanol to form a B 6 solution; mixing the solution A 6 and the solution B 6 according to any proportion of 9:1-0:10 to form a color developing layer solution, soaking filter paper in the color developing layer solution, drying to form color developing layer base paper, attaching the reaction layer base paper to the color developing layer base paper to form amine base paper, or dripping 5-30ul of the color developing layer solution into holes of the punched filter paper, drying to form amine test paper color developing holes, and cutting the reaction layer base paper into proper sizes and attaching the color developing holes to form amine reaction holes.
The test method adopting the vaginal secretion multi-connected test paper comprises the following steps of:
rotating the aseptic cotton swab or the aseptic swab at a fornix behind the vagina for 10-20 seconds until secretion is clearly observed to be attached to the cotton swab or the aseptic swab;
collecting samples by using sterile swabs or sterile swabs, putting the samples into soft test tubes, adding physiological saline into each sample, repeatedly rinsing, and fully eluting the samples to obtain sample liquid;
taking out the dry chemical test paper 10, and dripping 20-30ul of samples into each project test paper hole, wherein about 1 drop of samples is dripped;
The test paper is placed on a specific instrument for detection, or after the test paper is incubated for 5 minutes at 37+/-1 ℃, the result is interpreted by comparing with the color code card 20 in 1 minute.
In the present embodiment, in the case of the present embodiment,
The detection principle of each item is as follows:
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items are compared to the color chart 20:
Leukocyte esterase:
the color of no color (-) or light purple (+ -) is negative, which indicates normal; mauve (+1- +3) is positive and the shade of the color indicates that the patient has different degrees of inflammatory response in the vagina.
Hydrogen peroxide:
The red or purplish red is negative (-), which indicates that a large amount of lactobacillus possibly exists and the vaginal flora is normal;
The bacterium is light red (+ -) and weak positive, which suggests that a moderate amount of lactobacillus exists, and that the vagina bacteria starts to show abnormal trend or is in the recovery period, and clinical re-judgment is needed to be combined, and the bacterium is usually judged to be negative; the non-chromogenic or yellowish positive (+) indicates that the vaginal flora is deregulated, and the vaginal environment is sick or in a sub-health state.
Sialidase:
The color is not developed or the yellow is negative (-); red or light purple is weakly positive (+ -), suggesting a possibility of bacterial vaginosis infection; redness or mauve positive (+), indicating a potential infection with Bacterial Vaginosis (BV).
N-acetylglucosaminidase:
The color is not developed or the light yellow is negative (-), which indicates normal; orange is weak positive (+ -), orange red is positive (+), weak positive or positive indicates infection by mold such as candida or trichomonas;
proline aminopeptidase:
The color is not developed or the light yellow is negative (-), which indicates normal; pale yellow is weakly positive (+ -), yellowish or purple yellow is positive (+), weakly positive or positive indicates a potential infection with Bacterial Vaginosis (BV).
Beta-glucuronidase:
the color is not developed as negative (-), and normal is indicated; red or light purple is weak positive (+ -), red or purple is positive (+), and positive or weak positive indicates that the vagina is infected with aerobic bacteria, and the vagina environment is sick or in sub-health state;
Coagulase:
The color is not developed or the light yellow color is negative (-), which indicates normal; pale yellow as weak positive (+ -); yellow or purple yellow positive (+) and positive or weak positive represent infection by aerobic bacteria such as staphylococcus aureus and streptococcus;
Lactic acid:
The red or purplish red is negative (-), which indicates that a large amount of lactobacillus possibly exists and the vaginal flora is normal;
The bacterium is light red (+ -) and weak positive, which suggests that a moderate amount of lactobacillus exists, and that the vagina bacteria starts to show abnormal trend or is in the recovery period, and clinical re-judgment is needed to be combined, and the bacterium is usually judged to be negative; the non-chromogenic or yellowish positive (+) indicates that the vaginal flora is deregulated, and the vaginal environment is sick or in a sub-health state.
Oxidase:
The color is not developed or the light yellow is negative (-), which indicates normal; bluish with weak positive (+ -); blue or bluish purple positive (+) positive or weak positive indicates a potential gonococcal infection;
Amine:
yellow negative (-) indicates normal; pale green as weak positive (+ -); green, blue-green, blue positive (+), positive or weak positive indicates a possible bacterial vaginosis.
The application of the vaginal secretion multi-joint detection test paper in detecting at least one of PH value, beneficial bacteria, leucocyte, lactobacillus, BV pathogenic bacteria, gonococcus, gardnerella and campylobacter, candida albicans and trichomonas, escherichia coli and group B streptococcus, staphylococcus aureus and enterococcus faecalis and leucorrhea in vaginal secretion.
The above disclosure is only a preferred embodiment of the present invention, and it should be understood that the scope of the invention is not limited thereto, and those skilled in the art will appreciate that all or part of the procedures described above can be performed according to the equivalent changes of the claims, and still fall within the scope of the present invention.
Claims (6)
1. A vaginal secretion multiplex test paper, comprising:
The dry chemical detection test paper consists of a plastic base material and project base paper/project reaction blocks, wherein the project base paper/project reaction blocks are pH value base paper/reaction blocks, hydrogen peroxide base paper/reaction blocks, leucocyte zymogen paper/reaction blocks, lactic acid base paper/reaction blocks, sialidase zymogen paper/reaction blocks, proline aminopeptidase paper/reaction blocks, N-acetaminoglycosidase zymogen paper/reaction blocks, beta-glucuronidase zymogen paper/reaction blocks, solidification zymogen paper/reaction blocks, oxidation zymogen paper/reaction blocks and amine base paper/reaction blocks, and the plastic base material is one of a base strip or a test paper card;
The color code card is combined with the dry chemical detection test paper;
the dry chemical test paper comprises a clamping type test paper and a strip type test paper, each prepared base paper is divided into test paper strips with certain specification according to the requirement, the test paper strips are adhered to the matrix strips, and the strip type test paper is manufactured after cutting; or cutting each item base paper according to the round hole size of the test paper card, and punching the cut item base paper into a corresponding round hole and preparing a card type detection test paper by referring to a corresponding item reaction block;
The leucocyte zymogen paper/reaction block comprises 0.5 g/L-1.0 g/L of a first reaction substrate, 0.1 g/L-1.0 g/L of a diazonium salt first color reagent, 1.0 g/L-10 g/L of a first reaction promoter, 0.5 g/L-1.0 g/L of a first stabilizer and PBS buffer solution, wherein the first reaction substrate is at least one of indoxyl esters and derivatives thereof, pyrrole esters and pyrrolyl amino acid ester derivatives thereof, the first stabilizer is at least one of beta-cyclodextrin, mannitol, ethylene glycol, glycerol, ethylenediamine tetraacetic acid sodium salt and ethylenediamine tetraacetic acid potassium salt, and the first reaction promoter is at least one of sodium chloride and potassium thiocyanate;
the hydrogen peroxide base paper/reaction block comprises 0.5 g/L-1.0 g/L of peroxidase or horseradish peroxidase, 0.1 g/L-0.6 g/L of second color development agent, 0.1 g/L-1.0 g/L of second reaction accelerator, and a second buffer solution as a solvent, wherein the second color development agent is as follows: at least one of 4-aminoantipyrine, DHBS and 1, 5-dimethyl-2-phenyl-4-amino-3-pyrazolone (4-AAP), wherein the second reaction promoter is at least one of N-ethyl-N- (2-hydroxy-3-sulfopropyl) m-toluidine and 3, 5-dichloro-2-hydroxybenzenesulphonic acid, and the second buffer solution is any one of PBS buffer, citric acid-trisodium citrate buffer and boric acid-borax buffer;
The pH value base paper/reaction block comprises 0.1-0.6 g/L bromocresol green, 0.1-2 g/L surfactant and 0.1-2 g/L second stabilizer, wherein the solvent is ethanol with the concentration of 10-100%, and the second stabilizer is at least one of sodium alginate, trehalose, sodium carboxymethylcellulose, polyvinylpyrrolidone and gelatin;
The beta-glucuronide zymogen paper/reaction block comprises a second reaction substrate composed of at least one of 0.1 g/L-4.0 g/L of 5-bromo-4-chloro-3-indolyl-beta-D-glucuronide sodium salt, 4-nitrophenyl-beta-D-glucopyranoside, 5-bromo-4-chloro-3-indolyl-beta-D-glucopyranoside and phenolphthalein glucuronide, 0.1 g/L-4.0 g/L of diazonium salt first color reagent, 0.2 g/L-6.0 g/L of third reaction promoter, 1 g/L-10 g/L of third stabilizing agent, the solvent is a third buffer solution, the third buffer solution is any one of PBS buffer solution, citric acid-trisodium citrate buffer solution, citric acid-TRIS buffer solution, boric acid-borax buffer solution and sodium acetate buffer solution, the third reaction promoter is at least one of magnesium chloride, zinc chloride, calcium chloride, ethylene glycol, sodium alginate, sodium salt of at least one of diamine, ethylene acetate, sodium salt of ethylene diamine and ethylene acetate, and at least one of stabilizing agents;
The zymogen paper/reaction block comprises glycyl-arginyl-4-methoxy-beta-naphthylamine with the concentration of 1 g/L-10 g/L as a third reaction substrate, diazonium salt type first color developing agent, fourth stabilizer which is a mixed solution of trehalose and sucrose, and solvent which is hydrochloric acid-TRIS buffer solution;
The N-acetylglucosaminidase paper/reaction block comprises a fourth reaction substrate composed of at least one of 0.1g/L-4g/L of 4-nitrophenyl-2-acetamido-2-deoxy-beta-D-glucopyranoside, 5-bromo-4-chloro-3-indolyl-beta-D-glucuronide, p-nitrophenol-N-acetyl-beta-D-aminoglycoside, 2-chloro-4-nitrophenyl-N-acetyl-beta-D-aminoglycoside, 4-nitrophenyl-N-acetyl-beta-D-aminoglycoside and 5- [4- (3-methoxy-benzalkene-rhodanine) ] -3-ammonium acetate-N-acetamido-beta-D-glucoside, the solvent is PBS buffer solution, the fourth reaction promoter is at least one of sodium tungstate and bovine serum albumin, and the fifth stabilizer is at least one of vitamin C, ascorbyl oxidase, glutathione and potassium iodide;
The zymolyside paper/reaction block comprises a chromogenic substrate layer and a chromogenic promoting layer, wherein the chromogenic substrate layer comprises 0.5 g/L-4.0 g/L of 5-bromo-4-chloro-3-indole-alpha-D-N-acetylneuraminic acid sodium salt, cresol blue-alpha-N-acetylneuraminic acid glycoside sodium salt, nitrobenzene-N-acetyl-alpha-sialidamide, naphthol-N-acetyl-alpha-sialidamide or 5-bromo-4 chloro-3-indolylacetylceramic acid salt, the chromogenic substrate layer comprises 0.5 g/L-1.0 g/L of diazonium salt, the fifth reaction promoting layer comprises 0.1 g/L-10 g/L of sodium tungstate, polyvinylpyrrolidone and at least one of thiols, and the sixth stabilizer comprises at least one of beta-cyclodextrin, mannitol, ethylene glycol, glycerol, ethylene diamine tetraacetic acid sodium salt, ethylene diamine tetraacetic acid potassium salt and bovine serum albumin;
The proline aminopeptidase paper/reaction block comprises a sixth reaction substrate prepared from at least one of L-proline-paranitroaniline trifluoroacetate, Z-glycylproline-paranitroaniline, L-proline beta-naphthamide hydrochloride and L-prolyl-paranitroaniline with the concentration of 0.4 g-5 g/L, and the solvent is a third buffer solution; the sixth reaction promoter is diazonium salt reagent, and the seventh stabilizer is at least one of tartaric acid, oxalic acid tetraacetic acid sodium salt, potassium salt and maleic acid;
The lactic acid base paper/reaction block comprises 0.1-1.0 g/L peroxidase, 0.1-1.0 g/L lactic acid oxidase, a surfactant, a third color developing agent which is 3,3', 5' -tetramethyl benzidine and a solvent which is PBS buffer solution;
The oxidation zymogen paper/reaction block comprises a seventh reaction substrate of 1 g/L-10 g/L tetramethyl p-phenylenediamine, 0.5 g/L-5 g/L surfactant and a second buffer solution as a solvent;
The amine base paper/reaction block consists of a reaction layer and a color development layer, wherein the reaction layer is a potassium hydroxide solution with the mass fraction of 0.1% -5%; the color-developing layer contains 20 mg/L-500 mg/L bromocresol green, 0.8 g/L-2 g/L surfactant and 10% -100% ethanol as solvent.
2. The vaginal secretion multi-linked test paper of claim 1, wherein the surfactant is at least one of OP emulsifier, triton X-100, tween 20, tween 40, tween 60, tween 80 and polyethylene glycol.
3. The preparation method of the vaginal secretion multi-joint detection test paper according to claim 1 is characterized by comprising the following steps of:
preparation of matrix strips or test paper cards:
adhering the project base paper on a sheet or a coiled material made of at least one material of PET, PVC, PP, PS or ABS to form a matrix strip; the test paper card is made of at least one material of ABS, PP, modified PP or PMMA, a plurality of round holes with diameter of 3 mm-7 mm are formed in the test paper card, the number of the round holes is consistent with that of the project reaction blocks, and each round hole is filled with the corresponding project reaction block;
Preparation of project base paper/project reaction block:
Preparing a leucocyte zymogen paper/reaction block, dissolving a first reaction substrate in a first PBS buffer solution with the pH value of 6-9 and the concentration of 0.1mM-2mM, adding 1g/L-10g/L of a first stabilizer, forming a solution A 1 by 0.5g/L-5g/L of a first reaction promoter, dissolving a diazonium salt type first color reagent in an organic acid solvent containing 0.5g/L-10g/L and adding 0.1g/L-5g/L of a surfactant to form a solution B 1, mixing the solution A 1 and the solution B 1 according to the volume ratio of 1:1-9:1 to form a leucocyte esterase solution, soaking filter paper in the leucocyte esterase solution, and drying to prepare the leucocyte enzyme paper block, or dripping 5-30 mu L of the leucocyte solution into a reaction hole of the punched filter paper, and drying to form a leucocyte esterase reaction hole;
Preparing hydrogen peroxide base paper/reaction block, dissolving peroxidase or horseradish peroxidase in a second buffer solution with a pH value of 5-8 and a concentration of 0.1mM-2mM, adding 0.1 g/L-0.6 g/L of a second color reagent and 0.1 g/L-1.0 g/L of a second reaction accelerator to prepare a hydrogen peroxide test solution, soaking filter paper in the hydrogen peroxide solution, and drying, or dripping 5-30 mu L of the hydrogen peroxide solution into a reaction hole of the punched filter paper, and drying to form a hydrogen peroxide reaction hole;
preparing pH value base paper/reaction block, dissolving bromocresol green and a surfactant in a 10% -100% ethanol solvent to form A 2 liquid, dissolving a second stabilizer in purified water or ethanol to form B 2 liquid, mixing the A 2 liquid and the B 2 liquid according to any volume ratio of 9:1-1:9 to form pH value solution, soaking filter paper in the pH value solution, and drying to obtain the pH value paper block, or dripping 5-30 mu l of the pH value solution into a reaction hole of the punched filter paper, and drying to form a pH value reaction hole;
Preparing beta-glucuronidase paper/reaction block, dissolving a second reaction substrate in a third buffer solution with the pH value of 4-9 and the concentration of 0.5-10 mM, adding 1g/L-10g/L of a third stabilizer and 0.5g/L-5g/L of a third reaction accelerator to form A 3 liquid, dissolving diazonium salt type first color reagent in an organic acid solution with the concentration of 0.5g/L-10g/L to form B 3 liquid, mixing the A 3 liquid and the B 3 liquid according to the volume ratio of 1:1 to form beta-glucuronidase solution, soaking filter paper in the beta-glucuronidase solution, and drying to prepare the beta-glucuronidase paper block, or dripping 5-30 mu L of the beta-glucuronidase solution into a reaction hole of the punched filter paper, and drying to form the beta-glucuronidase reaction hole;
preparing a clotting zymogen paper/reaction block, namely dissolving a third reaction substrate in a hydrochloric acid-TRIS buffer solution with the pH value of 3-7 and the concentration of 0.1-5 mM, buffering the solution, adding a fourth stabilizer, dissolving the solution to form A 4 solution, dissolving diazonium salt type first color reagent in an organic acid solution with the concentration of 0.5-10 g/L to form B 4 solution, soaking and drying filter paper in a clotting enzyme A 4 solution, soaking the filter paper in a clotting enzyme B 4 solution, and drying to obtain a clotting enzyme paper block, or dripping 5-30 mu L of clotting enzyme A 4 solution into a reaction hole of the punched filter paper, drying and dripping 5-30 mu L of clotting enzyme A 4 solution, and drying to obtain a clotting enzyme reaction hole;
Preparing N-acetylglucosaminidase paper/reaction block, dissolving 0.1g/L-4g/L of fourth reaction substrate in PBS buffer solution with pH value of 7-10 and concentration of 0.1mM-10mM, adding 1g/L-10g/L of fifth stabilizer and 0.5g/L-5g/L of fourth reaction promoter to form N-acetylglucosaminidase solution, soaking filter paper in the N-acetylglucosaminidase solution, and drying to obtain N-acetylglucosaminidase paper block, or dripping 5-30 mu L of N-acetylglucosaminidase solution into a reaction hole of the punched filter paper, and drying to form N-acetylglucosaminidase reaction hole;
Preparing a sialidase paper/reaction block, namely dissolving 0.5g/L-4.0g/L of a fifth reaction substrate in a buffer solution with the pH value of 3-7 and the concentration of 0.1mM-10mM, adding 1g/L-10g/L of a sixth stabilizer, dissolving 0.1g/L-10g/L of a fifth reaction promoter into a solution A 5, dissolving diazonium salt type first color reagent in an organic acid solution containing 0.1g/L-10g/L to form a solution B 5, mixing the solution A 5 and the solution B 5 according to the volume ratio of 1:1 to form a sialidase solution, soaking filter paper in the sialidase solution, and drying to prepare the sialidase solution or dripping 5-30 mu L of the sialidase solution into a reaction hole of the punched filter paper, and drying to form a sialidase reaction hole;
Preparing proline aminopeptidase paper/reaction block, namely dissolving 0.4g/L-5g/L of sixth reaction substrate in a buffer solution with the pH value of 6-8 and the concentration of 0.1mM-10mM, adding 1g/L-10g/L of seventh stabilizer and 0.5g/L-5g/L of sixth reaction accelerator to form proline aminopeptidase solution, soaking filter paper in the proline aminopeptidase solution, and drying to obtain the proline aminopeptidase paper block, or dripping 5-30 mu L of proline aminopeptidase solution into a reaction hole of the punched filter paper, and drying to form a proline aminopeptidase reaction hole;
Preparing lactic acid base paper/reaction block, dissolving 0.1-1.0 g/L peroxidase, 0.1-1.0 g/L lactic acid oxidase, surfactant and 3,3', 5' -tetramethyl benzidine as a third color reagent in a first PBS buffer solution with pH value of 5-7 to form lactic acid solution, soaking filter paper in the lactic acid solution and drying to obtain lactic acid paper block, or dripping 5-30 mu L of lactic acid solution into a reaction hole of the punched filter paper and drying to form lactic acid reaction hole;
preparing oxidation zymogen paper/reaction block, dissolving 1 g/L-10 g/L of seventh reaction substrate in a second buffer solution, adding 0.5 g/L-5 g/L of surfactant to form oxidase reagent solution, soaking filter paper in the oxidase solution, and drying to obtain the product, or dripping 5-30 mu L of oxidase solution into a reaction hole of the punched filter paper, and drying to form an oxidase reaction hole;
Preparing amine base paper/reaction block, soaking and drying 0.1% -5% potassium hydroxide solution and filter paper to form reaction layer base paper; dissolving bromocresol green and a surfactant in a 10% -100% ethanol solvent to form A 6 liquid; dissolving a second stabilizer in purified water or ethanol to form a B 6 solution; mixing the solution A 6 and the solution B 6 according to any ratio of 9:1 to form a chromogenic layer solution, soaking filter paper in the chromogenic layer solution, drying to form chromogenic layer base paper, attaching the reaction layer base paper on the chromogenic layer base paper to form amine base paper, or dripping 5-30 mu l of the chromogenic layer solution into holes of the punched filter paper, drying to form amine test paper chromogenic holes, dividing the reaction layer base paper into proper sizes, and attaching the proper sizes of the reaction layer base paper on the chromogenic holes to form amine reaction holes.
4. A vaginal discharge multiple test strip as claimed in claim 3 wherein said organic acid is at least one of sodium benzoate, benzoic acid, citric acid and tartaric acid in the range of 0.1g/L to 10 g/L.
5. A test method using a vaginal discharge multiplex test strip as claimed in claim 3, comprising the steps of:
rotating the aseptic cotton swab or the aseptic swab at a fornix behind the vagina for 10-20 seconds until secretion is clearly observed to be attached to the cotton swab or the aseptic swab;
collecting samples by using sterile swabs or sterile swabs, putting the samples into soft test tubes, adding physiological saline into each sample, repeatedly rinsing, and fully eluting the samples to obtain sample liquid;
taking out the dry chemical detection test paper, and dripping 20-30 mu l of sample and about 1 drop of sample into each test paper hole of each project;
The test strip is placed on an instrument for detection, or after the test strip is incubated for 5 minutes at 37+/-1 ℃, the result is interpreted by comparing with a color code card within 1 minute.
6. Use of a vaginal secretion test strip as claimed in any one of claims 1, 3 and 5 for detecting at least one of pH, white blood cells, lactobacillus, gonococcus, gardnerella and campylobacter, candida albicans and trichomonas, escherichia coli and staphylococcus aureus in vaginal secretions.
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| CN114280040A (en) * | 2021-11-18 | 2022-04-05 | 中国农业科学院油料作物研究所 | Test strip for detecting peroxide value of edible oil, preparation method and application thereof |
| CN114235789A (en) * | 2021-11-29 | 2022-03-25 | 北京华晟源医疗科技有限公司 | Preparation method of beta-glucuronidase detection paper |
| CN114317678B (en) * | 2021-12-31 | 2023-11-10 | 港龙生物技术（深圳）有限公司 | Biological paper chip, high-throughput multi-detection microplate device, preparation method and kit for vaginal inflammation multi-detection |
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