CN114441523A - Preparation method of kit for detecting sulfhydryl metabolites in urine - Google Patents
Preparation method of kit for detecting sulfhydryl metabolites in urine Download PDFInfo
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- CN114441523A CN114441523A CN202210123639.3A CN202210123639A CN114441523A CN 114441523 A CN114441523 A CN 114441523A CN 202210123639 A CN202210123639 A CN 202210123639A CN 114441523 A CN114441523 A CN 114441523A
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- 238000002360 preparation method Methods 0.000 title claims abstract description 12
- 210000002700 urine Anatomy 0.000 title claims abstract description 10
- 125000003396 thiol group Chemical group [H]S* 0.000 title claims description 8
- 239000000243 solution Substances 0.000 claims abstract description 62
- 239000003153 chemical reaction reagent Substances 0.000 claims abstract description 40
- 238000012360 testing method Methods 0.000 claims abstract description 31
- 239000012153 distilled water Substances 0.000 claims abstract description 19
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical group O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 19
- 239000007853 buffer solution Substances 0.000 claims abstract description 11
- IYDGMDWEHDFVQI-UHFFFAOYSA-N phosphoric acid;trioxotungsten Chemical compound O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.OP(O)(O)=O IYDGMDWEHDFVQI-UHFFFAOYSA-N 0.000 claims abstract description 9
- 239000012445 acidic reagent Substances 0.000 claims abstract description 8
- 238000011981 development test Methods 0.000 claims abstract description 7
- 238000011161 development Methods 0.000 claims abstract description 6
- 229960002523 mercuric chloride Drugs 0.000 claims abstract description 6
- LWJROJCJINYWOX-UHFFFAOYSA-L mercury dichloride Chemical group Cl[Hg]Cl LWJROJCJINYWOX-UHFFFAOYSA-L 0.000 claims abstract description 6
- 238000001035 drying Methods 0.000 claims abstract description 5
- DOBUSJIVSSJEDA-UHFFFAOYSA-L 1,3-dioxa-2$l^{6}-thia-4-mercuracyclobutane 2,2-dioxide Chemical compound [Hg+2].[O-]S([O-])(=O)=O DOBUSJIVSSJEDA-UHFFFAOYSA-L 0.000 claims abstract description 4
- 229940074994 mercuric sulfate Drugs 0.000 claims abstract description 4
- 229910000372 mercury(II) sulfate Inorganic materials 0.000 claims abstract description 4
- 238000002156 mixing Methods 0.000 claims abstract description 3
- 238000004806 packaging method and process Methods 0.000 claims abstract 3
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 claims description 10
- XMVONEAAOPAGAO-UHFFFAOYSA-N sodium tungstate Chemical compound [Na+].[Na+].[O-][W]([O-])(=O)=O XMVONEAAOPAGAO-UHFFFAOYSA-N 0.000 claims description 8
- 239000003795 chemical substances by application Substances 0.000 claims description 7
- FHVDTGUDJYJELY-UHFFFAOYSA-N 6-{[2-carboxy-4,5-dihydroxy-6-(phosphanyloxy)oxan-3-yl]oxy}-4,5-dihydroxy-3-phosphanyloxane-2-carboxylic acid Chemical compound O1C(C(O)=O)C(P)C(O)C(O)C1OC1C(C(O)=O)OC(OP)C(O)C1O FHVDTGUDJYJELY-UHFFFAOYSA-N 0.000 claims description 5
- 229940072056 alginate Drugs 0.000 claims description 5
- 235000010443 alginic acid Nutrition 0.000 claims description 5
- 229920000615 alginic acid Polymers 0.000 claims description 5
- 229910000147 aluminium phosphate Inorganic materials 0.000 claims description 5
- 238000009835 boiling Methods 0.000 claims description 5
- 238000001816 cooling Methods 0.000 claims description 5
- RUTXIHLAWFEWGM-UHFFFAOYSA-H iron(3+) sulfate Chemical compound [Fe+3].[Fe+3].[O-]S([O-])(=O)=O.[O-]S([O-])(=O)=O.[O-]S([O-])(=O)=O RUTXIHLAWFEWGM-UHFFFAOYSA-H 0.000 claims description 5
- 229910000360 iron(III) sulfate Inorganic materials 0.000 claims description 5
- 235000015110 jellies Nutrition 0.000 claims description 5
- 239000008274 jelly Substances 0.000 claims description 5
- INHCSSUBVCNVSK-UHFFFAOYSA-L lithium sulfate Inorganic materials [Li+].[Li+].[O-]S([O-])(=O)=O INHCSSUBVCNVSK-UHFFFAOYSA-L 0.000 claims description 5
- 238000010992 reflux Methods 0.000 claims description 5
- RBTVSNLYYIMMKS-UHFFFAOYSA-N tert-butyl 3-aminoazetidine-1-carboxylate;hydrochloride Chemical compound Cl.CC(C)(C)OC(=O)N1CC(N)C1 RBTVSNLYYIMMKS-UHFFFAOYSA-N 0.000 claims description 5
- 238000005303 weighing Methods 0.000 claims description 5
- 239000008055 phosphate buffer solution Substances 0.000 claims description 4
- 239000008351 acetate buffer Substances 0.000 claims description 3
- 238000000034 method Methods 0.000 claims description 3
- 239000003708 ampul Substances 0.000 claims description 2
- 239000000203 mixture Substances 0.000 abstract description 10
- 206010008342 Cervix carcinoma Diseases 0.000 abstract description 5
- 208000006105 Uterine Cervical Neoplasms Diseases 0.000 abstract description 5
- 201000010881 cervical cancer Diseases 0.000 abstract description 5
- 238000001514 detection method Methods 0.000 abstract description 5
- 238000003745 diagnosis Methods 0.000 abstract description 2
- 239000011363 dried mixture Substances 0.000 abstract 1
- 239000002207 metabolite Substances 0.000 abstract 1
- 238000002791 soaking Methods 0.000 abstract 1
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 9
- BNIILDVGGAEEIG-UHFFFAOYSA-L disodium hydrogen phosphate Chemical compound [Na+].[Na+].OP([O-])([O-])=O BNIILDVGGAEEIG-UHFFFAOYSA-L 0.000 description 5
- 229910000403 monosodium phosphate Inorganic materials 0.000 description 5
- 235000019799 monosodium phosphate Nutrition 0.000 description 5
- AJPJDKMHJJGVTQ-UHFFFAOYSA-M sodium dihydrogen phosphate Chemical compound [Na+].OP(O)([O-])=O AJPJDKMHJJGVTQ-UHFFFAOYSA-M 0.000 description 5
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 3
- 238000011049 filling Methods 0.000 description 3
- 239000001632 sodium acetate Substances 0.000 description 3
- 235000017281 sodium acetate Nutrition 0.000 description 3
- 206010011409 Cross infection Diseases 0.000 description 2
- 241000701806 Human papillomavirus Species 0.000 description 2
- 206010029803 Nosocomial infection Diseases 0.000 description 2
- 238000012216 screening Methods 0.000 description 2
- 206010048038 Wound infection Diseases 0.000 description 1
- 239000003086 colorant Substances 0.000 description 1
- 208000014674 injury Diseases 0.000 description 1
- 230000003902 lesion Effects 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 238000009595 pap smear Methods 0.000 description 1
- 238000010827 pathological analysis Methods 0.000 description 1
- 238000005057 refrigeration Methods 0.000 description 1
- 230000008733 trauma Effects 0.000 description 1
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
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- Physics & Mathematics (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Plasma & Fusion (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Investigating Or Analyzing Non-Biological Materials By The Use Of Chemical Means (AREA)
Abstract
A preparation method of a kit for detecting sulfydryl metabolites in urine comprises a contrast reagent and a test reagent, and color development test paper, wherein the solution of the test reagent is distilled water, the solution of the contrast reagent is a mercuric chloride solution or a mercuric sulfate solution, and the color development test paper is prepared by uniformly mixing a phosphotungstic acid reagent solution and a buffer solution according to the volume ratio of 1:1, soaking the mixture in a color development reagent solution by using qualitative slow-speed filter paper, taking out the mixture, drying the mixture in a box for 1 hour, and packaging the dried mixture by using a plastic bag. The detection kit has the characteristics of high accuracy, good specificity and the like, and the positive coincidence rate is 87.4% by detecting 215 positive 188 cervical cancer patients with confirmed diagnosis; 385 cases of general population are screened by the reagent, positive 7 cases are screened, the positive rate is 1.8 percent, and the specificity is 98.2 percent; meanwhile, the same population is detected by the same reagents on the market, 27 positive cases are obtained, the positive rate is 7 percent, and the specificity is 93 percent.
Description
Technical Field
The invention relates to a preparation method of a detection kit, in particular to a preparation method of a kit for detecting sulfhydryl metabolites in urine.
Background
The obvious rise of the level of sulfhydryl metabolites in the urine of women can indicate the occurrence of cervical cancer lesion. The common cervical cancer screening methods at present comprise HPV (human papilloma virus) detection, TCT (liquid-based thin layer cell detection), pap smear detection and pathological diagnosis. These methods are relatively complex to operate, have low accuracy, and have certain trauma to the examined person and cross infection.
Disclosure of Invention
The invention aims to provide a preparation method of a kit for detecting sulfhydryl metabolites in urine, which is simple to operate and high in accuracy and can avoid wound and cross infection of a detected person.
The invention relates to a preparation method of a kit for detecting sulfhydryl metabolites in urine, which comprises a contrast reagent and a test reagent, wherein the contrast reagent and the test reagent are respectively packed in an ampoule, two pieces of same color development test paper are used, and the solution of the test reagent is distilled water, and the preparation method comprises the following specific steps:
the solution of the contrast reagent is a mercuric chloride solution or a mercuric sulfate solution, and the concentration is 0.2-0.5 mol/l.
Preparation of color test paper:
1. weighing 60-80 g of sodium tungstate, dissolving the sodium tungstate in 300-500 ml of distilled water, adding 30-50 ml of 85% phosphoric acid, refluxing and boiling for 2 hours, and then cooling to room temperature; adding 10g-20g of ferric sulfate, fully dissolving, and adding distilled water to a constant volume of 1000 ml; adding 30-50 g of lithium sulfate and 50-70 ml of 0.5% alginate jelly, and shaking up; refrigerating at 2-8 deg.c for further use;
2. the pH value of the buffer solution is 5.4-5.8, 1000ml of acetate buffer solution (sodium acetate solution/acetic acid solution) or 1000ml of phosphate buffer solution (sodium dihydrogen phosphate solution/disodium hydrogen phosphate solution) is prepared according to the requirement of the pH value, and the solution is refrigerated and stored at the temperature of 2-8 ℃ for later use;
uniformly mixing a phosphotungstic acid reagent solution and a buffer solution according to the volume ratio of 1:1 to obtain a color developing agent, and refrigerating and storing at the temperature of 2-8 ℃ for later use;
3. the color test paper is made into a circular test paper with the diameter of 1cm by using qualitative slow filter paper, is soaked in a color development agent solution for 3-5 minutes, is taken out and is dried in a thermostat at 26 ℃ for 1 hour, and is packaged by a plastic bag.
When the kit is used for the general survey/screening of cervical cancer, 0.2ml of morning urine is respectively taken and dripped into a control reagent bottle and a test reagent bottle of the kit, the test reagent bottles are uniformly shaken and stood for a moment, and then color developing test paper is respectively added. Observing the color change of the two pieces of test paper, and judging the test paper to be negative if the colors of the two pieces of test paper are consistent; if the two pieces of test paper are different in color and the color of the test side (blue) is darker than that of the control side, the test side is judged to be positive.
The kit for detecting the sulfhydryl metabolites in the urine has the characteristics of high accuracy, good specificity and the like, and the positive coincidence rate is 87.4 percent by detecting 215 positive 188 positive cervical cancer patients with confirmed diagnosis; 385 cases of general population are screened by the reagent, 7 cases of positive are screened, the positive rate is 1.8 percent, and the specificity is 98.2 percent; meanwhile, the same population is detected by the same reagent on the market, 27 positive cases are obtained, the positive rate is 7 percent, and the specificity is 93 percent.
Detailed Description
Example 1: (reagent solution is prepared according to the filling amount of each tube of the kit, 500 boxes of kit are made)
Firstly, preparing 100ml of distilled water by using distilled water as a solution of a test reagent;
secondly, preparing 100ml of 0.2mol/l mercuric chloride solution by taking the solution of the contrast reagent as the mercuric chloride solution;
and thirdly, the color developing agent is 2000ml of a mixture of phosphotungstic acid reagent solution and buffer solution according to the volume ratio of 1: 1.
1. Preparing a phosphotungstic acid reagent solution, weighing 61g of sodium tungstate, dissolving the sodium tungstate in 320ml of distilled water, adding 35ml of 85% phosphoric acid, refluxing and boiling for 2 hours, then cooling to room temperature, adding 15g of ferric sulfate, fully dissolving, and adding distilled water to fix the volume to 1000 ml. Finally, 35g of lithium sulfate and 50ml of 0.5 percent alginate jelly are added, evenly shaken and stored at the temperature of 2-8 ℃ for standby.
2. The pH of the buffer solution is 5.4, 1000ml of acetate buffer solution (sodium acetate solution/acetic acid solution) is prepared according to the requirement of the pH value, 833.3ml of 2mol/l sodium acetate solution is mixed with 166.7ml of 2mol/l acetic acid solution, and the mixture is refrigerated and stored at the temperature of 2-8 ℃ for standby.
The solutions prepared from the above "one" to "three" are respectively dispensed into 1ml ampoules according to 1 × 0.2ml of the control reagent and 1 × 0.2ml of the test reagent; 1000 pieces of color development test paper (circular test paper with the diameter of 1cm and made by qualitative slow-speed filter paper) are soaked in the color development reagent solution for 3-5 minutes, taken out and placed in a thermostat at 26 ℃ for drying for 1 hour, and packaged by plastic bags, wherein 2 pieces are packed in each bag. A total of 500 kits can be made.
Example 2: (reagent solution is prepared according to the filling amount of each tube of the kit, 500 boxes of kit are made)
Firstly, preparing 100ml of distilled water by using distilled water as a solution of a test reagent;
and thirdly, the color developing agent is 2000ml of a mixture of phosphotungstic acid reagent solution and buffer solution according to the volume ratio of 1: 1.
1. Preparation of reagent solution such as phosphotungstic acid, and the like, weighing 65g of sodium tungstate, dissolving in 350ml of distilled water, adding 40ml of 85% phosphoric acid, refluxing and boiling for 2 hours, then cooling to room temperature, adding 20g of ferric sulfate, fully dissolving, and adding distilled water to fix the volume to 1000 ml. Finally, 35g of lithium sulfate and 55ml of 0.5 percent alginate jelly are added, evenly shaken and stored at the temperature of 2-8 ℃ for standby.
2. The pH value of the buffer solution is 5.7, 1000ml of phosphate buffer solution (sodium dihydrogen phosphate solution/disodium hydrogen phosphate solution) is prepared according to the pH value requirement, 0.2mol/l of sodium dihydrogen phosphate solution is 935ml and 0.2mol/l of disodium hydrogen phosphate solution is added to 65ml and mixed, and the mixture is refrigerated and stored at the temperature of 2-8 ℃ for standby.
The solutions prepared from the above "one" to "three" are respectively dispensed into 1ml ampoules according to 1 × 0.2ml of the control reagent and 1 × 0.2ml of the test reagent; 1000 pieces of color development test paper (circular test paper with the diameter of 1cm and made by qualitative slow-speed filter paper) are soaked in the color development reagent solution for 3-5 minutes, taken out and placed in a thermostat at 26 ℃ for drying for 1 hour, and packaged by plastic bags, wherein 2 pieces are packed in each bag. A total of 500 kits can be made.
Example 3: (reagent solution is prepared according to the proportion of the tube filling amount of the kit, 1000 boxes of kit are made)
Firstly, preparing 200ml of distilled water by using a solution of a test reagent as the distilled water;
secondly, preparing 200ml of 0.5mol/l mercuric sulfate solution by using a solution of a contrast reagent as a mercuric chloride solution;
and thirdly, the color developing agent is a mixture of phosphotungstic acid reagent solution and buffer solution in a volume ratio of 1:1, and the volume of the mixture is 4000 ml.
1. Preparing a phosphotungstic acid reagent solution, weighing 80g of sodium tungstate, dissolving in 700ml of distilled water, adding 80ml of 85% phosphoric acid, refluxing and boiling for 2 hours, then cooling to room temperature, adding 18g of ferric sulfate, fully dissolving, and adding distilled water to reach a constant volume of 2000 ml. Finally, 65g of lithium sulfate and 65ml of 0.5 percent alginate jelly are added, evenly shaken and stored at the temperature of 2-8 ℃ for standby.
2. The pH of the buffer solution is 5.8, 2000ml of phosphate buffer solution (sodium dihydrogen phosphate solution/disodium hydrogen phosphate solution) is prepared according to the pH requirement, 0.2mol/l of sodium dihydrogen phosphate solution 1840ml is mixed with 0.2mol/l of disodium hydrogen phosphate solution 160ml, and the mixture is stored at the temperature of 2-8 ℃ for later use by refrigeration.
The solutions prepared from the above "one" to "three" are respectively dispensed into 1ml ampoules according to 1 × 0.2ml of the control reagent and 1 × 0.2ml of the test reagent; 2000 pieces of color test paper (circular test paper with the diameter of 1cm and made by qualitative slow-speed filter paper) are soaked in the color development agent solution for 3-5 minutes, taken out and placed in a thermostat at 26 ℃ for drying for 1 hour, and packaged by plastic bags, wherein 2 pieces are packed in each bag. A total of 1000 kits can be made.
Claims (1)
1. A preparation method of a kit for detecting sulfhydryl metabolites in urine comprises the steps of packaging a contrast reagent and a test reagent by an ampoule, packaging two same pieces of color development test paper, and using distilled water as a solution of the test reagent, and is characterized in that: the method comprises the following specific steps:
the solution of the contrast reagent is a mercuric chloride solution or a mercuric sulfate solution, and the concentration is 0.2-0.5 mol/l;
preparation of color test paper:
(1) weighing 60-80 g of sodium tungstate, dissolving the sodium tungstate in 300-500 ml of distilled water, adding 30-50 ml of 85% phosphoric acid, refluxing and boiling for 2 hours, and then cooling to room temperature; adding 10g-20g of ferric sulfate, fully dissolving, and adding distilled water to a constant volume of 1000 ml; adding 30-50 g of lithium sulfate and 50-70 ml of 0.5% alginate jelly, and shaking up; refrigerating at 2-8 deg.c for further use;
(2) preparing 1000ml of acetate buffer solution or 1000ml of phosphate buffer solution according to the pH value requirement, and refrigerating at 2-8 ℃ for later use, wherein the pH value of the buffer solution is 5.4-5.8;
uniformly mixing a phosphotungstic acid reagent solution and a buffer solution according to the volume ratio of 1:1 to obtain a color developing agent, and refrigerating and storing at the temperature of 2-8 ℃ for later use;
(3) and the color development test paper is made into a round test paper by using qualitative slow filter paper, is soaked in the color development reagent solution for 3-5 minutes, is taken out and is placed in a constant temperature box at 26 ℃ for drying for 1 hour, and is packaged by a plastic bag.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210123639.3A CN114441523A (en) | 2022-02-10 | 2022-02-10 | Preparation method of kit for detecting sulfhydryl metabolites in urine |
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| Application Number | Priority Date | Filing Date | Title |
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| CN202210123639.3A CN114441523A (en) | 2022-02-10 | 2022-02-10 | Preparation method of kit for detecting sulfhydryl metabolites in urine |
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| CN114441523A true CN114441523A (en) | 2022-05-06 |
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| CN202210123639.3A Pending CN114441523A (en) | 2022-02-10 | 2022-02-10 | Preparation method of kit for detecting sulfhydryl metabolites in urine |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115290904A (en) * | 2022-10-09 | 2022-11-04 | 山东三医生物技术有限公司 | In-situ hybridization HPV detection test strip, kit and preparation method thereof |
| CN115372605A (en) * | 2022-07-08 | 2022-11-22 | 北京常有生物科技有限公司 | Urine sulfhydryl compound detection reagent, detection test paper and preparation method of detection kit |
| CN116559448A (en) * | 2023-07-10 | 2023-08-08 | 山东三医生物技术有限公司 | Test paper for detecting HPV (human papilloma Virus) based on HPVC1 protein and preparation method thereof |
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| CN106468715A (en) * | 2016-06-21 | 2017-03-01 | 安徽康乐美生物科技有限公司 | A kind of cervical cancer urine detection reagent and its preparation method and application |
| CN110672593A (en) * | 2019-08-19 | 2020-01-10 | 杭州爱光医疗器械有限公司 | Sulfhydryl compound detection reagent, preparation method and kit thereof |
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Patent Citations (5)
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN115372605A (en) * | 2022-07-08 | 2022-11-22 | 北京常有生物科技有限公司 | Urine sulfhydryl compound detection reagent, detection test paper and preparation method of detection kit |
| CN115290904A (en) * | 2022-10-09 | 2022-11-04 | 山东三医生物技术有限公司 | In-situ hybridization HPV detection test strip, kit and preparation method thereof |
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Application publication date: 20220506 |