CN101487041A - Bacterial vaginopathy combined detection reagent - Google Patents
Bacterial vaginopathy combined detection reagent Download PDFInfo
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- CN101487041A CN101487041A CNA2009100374352A CN200910037435A CN101487041A CN 101487041 A CN101487041 A CN 101487041A CN A2009100374352 A CNA2009100374352 A CN A2009100374352A CN 200910037435 A CN200910037435 A CN 200910037435A CN 101487041 A CN101487041 A CN 101487041A
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Abstract
The invention discloses a bacterial vaginosis joint detecting reagent, which comprises a hydrogen peroxide reaction pad foam fluid, a neuraminidase reaction pad foam fluid, a leukocyte esterase reaction pad foam fluid and a pH reaction pad foam fluid. Compared with the prior art, the bacterial vaginosis joint detecting reagent has the advantages of simultaneous detection of multiple data sets, respective reflection of microecological environment, pathogenic bacteria and secretions situations, more comprehensive and reliable detection, simple and quick operation, no need of instrument and equipment, and lower cost.
Description
Technical field
The present invention relates to a kind of in-vitro diagnosis detection reagent, relate in particular to a kind of bacterial vaginopathy combined detection reagent, can estimate the pathogenic situation of microecology in vaginas environment, degree of cleaning and bacterial vaginosis by this detection reagent.
Background technology
Bacterial vaginosis (Bactarial Vaginosis, be called for short BV) be the modal vaginal infection diseases of the women of child-bearing age, domestic being reported in the healthy women, the BV morbidity is 18.9%, having among the patient of unusual vaginal secretions in gynaecology's outpatient service is 43.3%, in the random crowd of property is 36.7%, in pregnant woman's morbidity is 12.5%, it is because vagina normal microflora (containing lactobacillus) is replaced by the mixed bacterial that gardnerella vaginalis, anerobe, mycoplasma hominis constitute, and causes flora imbalance and the disease that causes.Though it does not belong to the venereal disease of domestic monitoring, merit attention that to be it see in that the venereal disease high risk population more more, but increase the susceptibility of venereal diseases such as acquired immune deficiency syndrome (AIDS), gonorrhoea, nongonococcal urethritis.The vaginal secretions that mainly shows as of BV increases with stench, and causes that uterine neck, body of uterus infect and the danger of inflammatory pelvic disease (PID).Because 40～50% BV patient does not have any symptom,, but to detect the chamber of experimentizing so the diagnosis of BV can not be only according to clinical manifestation.How finding BV virus people in time and taking the corresponding treatment measure early is the key point of diagnosis and treatment BV.
The common diagnostic means of bacterial vaginosis has methods such as traditional bacteriological detection, Amsel diagnostic method, ELISA (enzyme connects immunosorbent and measures) method, polymerase chain reaction method and zymochemistry reaction method.Because the bacteriology checking operation is cumbersome, seldom use.The Amsel diagnostic method is the gold standard of internationally recognized diagnosis BV, and three then diagnosable BV of being of positive performance are promptly arranged in following four: the first, and vagina pH〉4.5; The second, the amine test positive; The 3rd, cell picks up scent in the smear; The 4th, vaginal secretions is thinning foreign odor.PH measures with test paper method and measures, and artificial error in judgement is bigger, and the mensuration of amine test varies with each individual, reliability with whether have sex act relevant in the recent period, and taste is more unpleasant in the mensuration process, influence staff's working attitude, therefore, present domestic less promoting the use of.Some improved diagnostic reagents have also been extended according to the Amsel diagnostic method, as the patent No. is Chinese utility model patent " bacterial vaginitis the quick diagnosis reagent kit " (Granted publication number: CN2760549Y) of ZL200420111847.9, all ingredients and the unification of detection object are located in the test kit, are used fast with convenient; And for example application number is that (publication number: CN1793865A), this application detects the pH value to adopting pH reagent to 200510022508.2 Chinese patent application open " leukorrhea pH, amine test combined detection reagent ", and the amine detection reagent has been done improvement.
The detection method of enzymic activity aspect is relevant with the meta-bolites of distinctive microorganism in the vagina according to BV, reach the principle of diagnosis vagina pathology situation by the meta-bolites that detects distinctive microorganism in the vagina, yet the specificity of individual event enzyme assay diagnosis BV is relatively poor, and reliability is not high.
For this reason, easy to detect, quick and reliable bacterial vaginopathy combined detection reagent slowly is applied, the common combination that neuraminidase and leukocyte esterase combination, neuraminidase and Trimethylamine 99 combination, neuraminidase and hydrogen peroxide are arranged etc.Disclosed document such as application number are open " the combined test kit for bacterial vagina disease " (publication number: CN1850985A) of the Chinese patent application of 200610056827.X, adopt hydrogen peroxide agent, leukocyte esterase reagent and neuraminidase agent combination in this application, overcome not objective, the not science and the inaccurate deficiency of traditional single means diagnosis vaginosis.
Summary of the invention
Technical problem to be solved by this invention is that a kind of easy to detect, quick and reliable good bacterial vaginopathy combined detection reagent is provided in addition at the above-mentioned state of the art.
The present invention solves the problems of the technologies described above the technical scheme that is adopted: a kind of bacterial vaginopathy combined detection reagent, comprise hydroperoxidation pad bubble liquid, neuraminidase reacting pad bubble liquid, leukocyte esterase reacting pad bubble liquid and pH reacting pad bubble liquid, it is characterized in that
Described hydrogen oxide reacting pad bubble liquid comprises the iron protochloride of following component and proportioning: 0.1～0.5mM, the starch of 0.1～0.5mM, the 2-hydroxyl-3 of 10～50mg/L, 5-dichloro benzosulfonic acid sodium salt, 300～500U enzyme activity/L horseradish peroxidase, volume percent are 0.01% 2-thioglycol, buffering system is a 0.1M PB liquid, and this PB liquid pH is 4～5;
Described neuraminidase reacting pad bubble liquid comprises following component and proportioning: 5 ' of 0.1～1.0mM-bromo-2 '-chloro-indoles-3 '-oxygen sialic acid glycosides or 5-bromo-4-chloro-3-indyl-α-D-N-n acetylneuraminic acid n or thymolphthalein acetyl neuraminic acid, 3-(N-p-toluenesulfonyl-L-alanyl oxygen base)-indoles of 0.1～1mg/ml, the glucose of 0.1～1mg/ml, buffering system is the PB liquid of 0.1M, and this PB liquid pH is 5～6;
Described leukocyte esterase reacting pad bubble liquid comprises 3-(N-p-toluenesulfonyl-L-alanyl oxygen base)-5-phenylpyrrole of 1-ethanoyl-5-bromo-4-chloro-indole-3-acetic ester of following component and proportioning: 0.5～1.5mM, the acetate of 0.2～1.0mM-5-bromo-4-chloro-3-indolol ester, 0.3～1.5mg/ml, the glucose of 0.1～1mg/ml; buffering system is the PB liquid of 0.1M; this PB liquid pH is 6～7
Described pH reacting pad bubble liquid adopts molten mass percent 0.1%～0.5% tetrabromo-mcresolsulfonphthalein that is made into of sodium hydroxide solution of 0.02M.
Further, also comprise diluent, this diluent adopts 0.01M～0.1M pH 6.9PIPES damping fluid.
As preferably, the employed indicator of described neuraminidase reacting pad bubble liquid is 0.01M～0.05M sodium hydroxide.
The carrier of described each reacting pad can adopt at least a among the fiber filter paper Grade3, quantitative paper of Whatman3MM glass fiber filter paper, Whatman glass fiber filter paper, GF/A Whatman.
Compared with prior art, the invention has the advantages that: can measure multi-group data simultaneously, reflect micro-ecological environment, pathogenic bacterium and secretory product situation respectively, measure more comprehensively reliably, simple to operate quick simultaneously, need not to use plant and instrument, cost is lower.
Embodiment
Below in conjunction with embodiment the present invention is described in further detail.
Embodiment: the bacterial vaginopathy combined detection reagent in the present embodiment is placed in the test kit, specifically comprises hydroperoxidation pad bubble liquid, neuraminidase reacting pad bubble liquid, leukocyte esterase reacting pad bubble liquid, pH reacting pad bubble liquid, diluent and indicator.
Hydrogen oxide reacting pad bubble liquid comprises the iron protochloride of following component and proportioning: 0.1～0.5mM, the starch of 0.1～0.5mM, the 2-hydroxyl-3 of 10～50mg/L, 5-dichloro benzosulfonic acid sodium salt, 300～500U enzyme activity/L horseradish peroxidase, volume percent are 0.01% 2-thioglycol, buffering system is a 0.1M PB liquid, and this PB liquid pH is 4～5;
Neuraminidase reacting pad bubble liquid comprises following component and proportioning: 5 ' of 0.1～1.0mM-bromo-2 '-chloro-indoles-3 '-oxygen sialic acid glycosides or 5-bromo-4-chloro-3-indyl-α-D-N-n acetylneuraminic acid n or thymolphthalein acetyl neuraminic acid, 3-(N-p-toluenesulfonyl-L-alanyl oxygen base)-indoles of 0.1～1mg/ml, the glucose of 0.1～1mg/ml, buffering system is the PB liquid of 0.1M, and this PB liquid pH is 5～6;
Leukocyte esterase reacting pad bubble liquid comprises 3-(N-p-toluenesulfonyl-L-alanyl oxygen base)-5-phenylpyrrole of 1-ethanoyl-5-bromo-4-chloro-indole-3-acetic ester of following component and proportioning: 0.5～1.5mM, the acetate of 0.2～1.0mM-5-bromo-4-chloro-3-indolol ester, 0.3～1.5mg/ml, the glucose of 0.1～1mg/ml; buffering system is the PB liquid of 0.1M; this PB liquid pH is 6～7
PH reacting pad bubble liquid adopts molten mass percent 0.1%～0.5% tetrabromo-mcresolsulfonphthalein that is made into of sodium hydroxide solution of 0.02M.
Diluent adopts 0.01M～0.1M pH 6.9PIPES damping fluid.
The used indicator that uses of neuraminidase reacting pad bubble liquid is 0.01M～0.05M sodium hydroxide.
Optimum ratio is as follows:
Hydrogen oxide reacting pad bubble liquid comprises the iron protochloride of following component and proportioning: 0.3mM (every liter of mmole), the starch of 0.3mM, the 2-hydroxyl-3 of 30mg/L, 5-dichloro benzosulfonic acid sodium salt, 400U enzyme activity/L horseradish peroxidase, volume percent are 0.01% 2-thioglycol, buffering system is a 0.1M PB liquid, and this PB liquid pH is 4.5;
Neuraminidase reacting pad bubble liquid comprises following component and proportioning: 5 ' of 0.5mM-bromo-2 '-chloro-indoles-3 '-oxygen sialic acid glycosides or 5-bromo-4-chloro-3-indyl-α-D-N-n acetylneuraminic acid n or thymolphthalein acetyl neuraminic acid, 3-(N-p-toluenesulfonyl-L-alanyl oxygen base)-indoles of 0.5mg/ml, the glucose of 0.5mg/ml, buffering system is the PB liquid of 0.1M, and this PB liquid pH is 5.5;
Leukocyte esterase reacting pad bubble liquid comprises 3-(N-p-toluenesulfonyl-L-alanyl oxygen base)-5-phenylpyrrole of 1-ethanoyl-5-bromo-4-chloro-indole-3-acetic ester of following component and proportioning: 1mM, the acetate of 0.6mM-5-bromo-4-chloro-3-indolol ester, 0.9mg/ml, the glucose of 0.5mg/ml; buffering system is the PB liquid of 0.1M; this PB liquid pH is 6.5
PH reacting pad bubble liquid adopts molten mass percent 0.3% tetrabromo-mcresolsulfonphthalein that is made into of sodium hydroxide solution of 0.02M.
Diluent adopts 0.05M pH 6.9PIPES damping fluid.
The used indicator that uses of neuraminidase reacting pad bubble liquid is 0.03M sodium hydroxide.
The carrier of each reacting pad can adopt fiber filter paper Grade3 and the common quantitative paper of Whatman3MM glass fiber filter paper, Whatman glass fiber filter paper, GF/A Whatman.
Get vaginal secretions and detect, directly judge ill situation by colour-change.Vaginopathy combined detection strip includes hydrogen peroxide H
2O
2, neuraminidase NAcase, leukocyte esterase LE and pH substrate reacting pad, H in sample
2O
2, neuraminidase NAcase, leukocyte esterase LE reach finite concentration, pH value greater than 4.5 o'clock, can with substrate generation chemistry or hydrolysis reaction separately, and make hydrogen peroxide H
2O
2Reacting pad is by colourless redness or the red-purple of becoming, neuraminidase NAcase reacting pad is by colourless redness, red-purple or the blueness of becoming, leukocyte esterase LE reacting pad is by colourless blue-greenish colour, blueness or the mazarine of becoming, and the pH reacting pad is by light blue blueness or the mazarine of becoming.When there not being H
2O
2, neuraminidase NAcase, leukocyte esterase LE or do not reach finite concentration, contact with substrate and above chemistry or hydrolysis reaction do not take place and keep original color, the pH value becomes yellow less than 4.5 o'clock (normally) reacting pads.
Operating process is as follows:
1. with test kit from 4～30 ℃ of taking-ups, equilibrate to room temperature and bring into use;
2. tear aluminium foil packing, take out reaction unit and take aluminum foil strip gently off, the reaction unit lucifuge is placed, and finished using on the same day;
3. rotated 10～20 seconds in the posterior fornix place with cotton swab, there is secretory product to adhere on the cotton swab to be as the criterion with clear seeing, has the reaction blind hole mid point of pH to print in the reaction unit sign earlier, at once observations, add 400 μ l diluents then and push cotton swab repeatedly, sample is disengaged;
4. every hole drips a sample of handling in other three holes of reaction unit, and about 35 μ l were not advisable to have reacting hole 1/2, drips another " colour developing liquid " afterwards in " neuraminidase " reaction;
5. reaction unit is placed on room temperature and leaves standstill, hydrogen peroxide H
2O
2, index sentence read result in 5～30min such as neuraminidase NAcase, leukocyte esterase LE, pH value sentence read result in 1～5sec;
A) hydroperoxidation hole: take on a red color, purple is negative; Show colourless positive.
B) neuraminidase reacting hole: it is negative not develop the color; Show red, red-violet colour or blue positive, it is dense to show blue expression sialidase activity.
C) leukocyte esterase reacting hole: it is negative not develop the color.Blue-greenish colour is "+", blue is " +++" for " ++ ", mazarine.
D) pH reacting hole: displaing yellow is negative, and is blue positive.
Claims (4)
1, a kind of bacterial vaginopathy combined detection reagent comprises hydroperoxidation pad bubble liquid, neuraminidase reacting pad bubble liquid, leukocyte esterase reacting pad bubble liquid and pH reacting pad bubble liquid, it is characterized in that
Described hydrogen oxide reacting pad bubble liquid comprises the iron protochloride of following component and proportioning: 0.1～0.5mM, the starch of 0.1～0.5mM, the 2-hydroxyl-3 of 10～50mg/L, 5-dichloro benzosulfonic acid sodium salt, 300～500U enzyme activity/L horseradish peroxidase, volume percent are 0.01% 2-thioglycol, buffering system is a 0.1M PB liquid, and this PB liquid pH is 4～5;
Described neuraminidase reacting pad bubble liquid comprises following component and proportioning: 5 ' of 0.1～1.0mM-bromo-2 '-chloro-indoles-3 '-oxygen sialic acid glycosides or 5-bromo-4-chloro-3-indyl-α-D-N-n acetylneuraminic acid n or thymolphthalein acetyl neuraminic acid, 3-(N-p-toluenesulfonyl-L-alanyl oxygen base)-indoles of 0.1～1mg/ml, the glucose of 0.1～1mg/ml, buffering system is the PB liquid of 0.1M, and this PB liquid pH is 5～6;
Described leukocyte esterase reacting pad bubble liquid comprises 3-(N-p-toluenesulfonyl-L-alanyl oxygen base)-5-phenylpyrrole of 1-ethanoyl-5-bromo-4-chloro-indole-3-acetic ester of following component and proportioning: 0.5～1.5mM, the acetate of 0.2～1.0mM-5-bromo-4-chloro-3-indolol ester, 0.3～1.5mg/ml, the glucose of 0.1～1mg/ml; buffering system is the PB liquid of 0.1M; this PB liquid pH is 6～7
Described pH reacting pad bubble liquid adopts molten mass percent 0.1%～0.5% tetrabromo-mcresolsulfonphthalein that is made into of sodium hydroxide solution of 0.02M.
2, bacterial vaginopathy combined detection reagent according to claim 1 is characterized in that also comprising diluent, and this diluent adopts 0.01M～0.1M pH6.9PIPES damping fluid.
3, bacterial vaginopathy combined detection reagent according to claim 1 is characterized in that the used indicator that uses of described neuraminidase reacting pad bubble liquid is 0.01M～0.05M sodium hydroxide.
4, bacterial vaginopathy combined detection reagent according to claim 1, the carrier that it is characterized in that described each reacting pad adopt at least a among the fiber filter paper Grade3, quantitative paper of Whatman3MM glass fiber filter paper, Whatman glass fiber filter paper, GF/A Whatman at least.
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| CNA2009100374352A CN101487041A (en) | 2009-02-26 | 2009-02-26 | Bacterial vaginopathy combined detection reagent |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA2009100374352A CN101487041A (en) | 2009-02-26 | 2009-02-26 | Bacterial vaginopathy combined detection reagent |
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| CNA2009100374352A Pending CN101487041A (en) | 2009-02-26 | 2009-02-26 | Bacterial vaginopathy combined detection reagent |
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| CN101975852A (en) * | 2010-09-16 | 2011-02-16 | 高翔 | Rapid detection device, detection test strip and detection method for bacterial vaginosis |
| CN102251022A (en) * | 2011-06-23 | 2011-11-23 | 泰普生物科学（中国）有限公司 | Rapid test kit for vaginitis |
| CN102253206A (en) * | 2011-06-23 | 2011-11-23 | 泰普生物科学（中国）有限公司 | Combined detection kit for vaginitis |
| CN102251021A (en) * | 2011-06-23 | 2011-11-23 | 泰普生物科学（中国）有限公司 | Detection kit for bacterial vaginosis (BV) and sialidase detection method |
| CN101792791B (en) * | 2009-12-30 | 2011-12-21 | 北京中生金域诊断技术有限公司 | Kit for detecting bacteria flora in vaginal secretion and preparation method thereof |
| CN102321730A (en) * | 2011-06-23 | 2012-01-18 | 泰普生物科学（中国）有限公司 | Joint vaginitis detection kit |
| CN102690861A (en) * | 2011-03-25 | 2012-09-26 | 安徽深蓝医疗科技有限公司 | Serial gynecological examination kit for bacterial vaginosis and application method thereof |
| US9034593B2 (en) | 2010-11-22 | 2015-05-19 | Kimberly-Clark Worldwide, Inc. | Vaginal indicator to detect biomarkers of good health |
| CN104911245A (en) * | 2014-03-13 | 2015-09-16 | 深圳市艾瑞生物科技有限公司 | Kit used for detecting bacterial vaginosis |
| CN108330159A (en) * | 2018-03-15 | 2018-07-27 | 深圳市瀚德标检生物工程有限公司 | A kind of detection reagent card and kit |
| CN108956827A (en) * | 2018-06-04 | 2018-12-07 | 长春百纯和成医药科技有限公司 | A kind of method HPLC method analysis and prepare 3- (N- p-toluenesulfonyl-L- alanyl oxygroup) indoles and its enantiomter |
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| CN109490519A (en) * | 2018-10-16 | 2019-03-19 | 迪瑞医疗科技股份有限公司 | A kind of leukocyte esterase Test paper and preparation method thereof |
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| CN101792791B (en) * | 2009-12-30 | 2011-12-21 | 北京中生金域诊断技术有限公司 | Kit for detecting bacteria flora in vaginal secretion and preparation method thereof |
| CN101975852A (en) * | 2010-09-16 | 2011-02-16 | 高翔 | Rapid detection device, detection test strip and detection method for bacterial vaginosis |
| US9034593B2 (en) | 2010-11-22 | 2015-05-19 | Kimberly-Clark Worldwide, Inc. | Vaginal indicator to detect biomarkers of good health |
| CN102690861A (en) * | 2011-03-25 | 2012-09-26 | 安徽深蓝医疗科技有限公司 | Serial gynecological examination kit for bacterial vaginosis and application method thereof |
| CN102251022A (en) * | 2011-06-23 | 2011-11-23 | 泰普生物科学（中国）有限公司 | Rapid test kit for vaginitis |
| CN102253206A (en) * | 2011-06-23 | 2011-11-23 | 泰普生物科学（中国）有限公司 | Combined detection kit for vaginitis |
| CN102251021A (en) * | 2011-06-23 | 2011-11-23 | 泰普生物科学（中国）有限公司 | Detection kit for bacterial vaginosis (BV) and sialidase detection method |
| CN102321730A (en) * | 2011-06-23 | 2012-01-18 | 泰普生物科学（中国）有限公司 | Joint vaginitis detection kit |
| CN104911245A (en) * | 2014-03-13 | 2015-09-16 | 深圳市艾瑞生物科技有限公司 | Kit used for detecting bacterial vaginosis |
| CN104911245B (en) * | 2014-03-13 | 2017-11-17 | 深圳市艾瑞生物科技有限公司 | A kind of kit for bacterial vaginosis BV detection |
| CN108330159A (en) * | 2018-03-15 | 2018-07-27 | 深圳市瀚德标检生物工程有限公司 | A kind of detection reagent card and kit |
| CN108956827A (en) * | 2018-06-04 | 2018-12-07 | 长春百纯和成医药科技有限公司 | A kind of method HPLC method analysis and prepare 3- (N- p-toluenesulfonyl-L- alanyl oxygroup) indoles and its enantiomter |
| CN109085255A (en) * | 2018-06-04 | 2018-12-25 | 长春百纯和成医药科技有限公司 | A kind of method HPLC method analysis and prepare 3- (N- p-toluenesulfonyl-L- alanyl oxygroup) -5- phenylpyrrole and its enantiomter |
| CN109085255B (en) * | 2018-06-04 | 2021-03-12 | 长春百纯和成医药科技有限公司 | Method for analyzing and preparing 3- (N-p-toluenesulfonyl-L-alanyloxy) -5-phenylpyrrole and enantiomer thereof by using HPLC method |
| CN108956827B (en) * | 2018-06-04 | 2021-05-04 | 长春百纯和成医药科技有限公司 | Method for analyzing and preparing 3- (N-p-toluenesulfonyl-L-alanyloxy) indole and enantiomer thereof by HPLC method |
| CN109490519A (en) * | 2018-10-16 | 2019-03-19 | 迪瑞医疗科技股份有限公司 | A kind of leukocyte esterase Test paper and preparation method thereof |
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