CN103865978B - Pseudocholinesterase detection kit and preparation method thereof - Google Patents
Pseudocholinesterase detection kit and preparation method thereof Download PDFInfo
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- CN103865978B CN103865978B CN201210529456.8A CN201210529456A CN103865978B CN 103865978 B CN103865978 B CN 103865978B CN 201210529456 A CN201210529456 A CN 201210529456A CN 103865978 B CN103865978 B CN 103865978B
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Abstract
The invention provides a kind of Pseudocholinesterase detection kit, this test kit, by following reagent 1 and reagent 2, forms in the ratio of 4 parts: 1 part: reagent 1:(1L)
; Reagent 2:(1L)
Description
Technical field
The present invention relates to biological reagent, be specifically related to a kind of detection kit, particularly relate to a kind of Pseudocholinesterase detection kit and preparation method thereof.
Background technology
Pseudocholinesterase (cholinesterase) is a class glycoprotein, is present in body with multiple isozyme form.Generally can be divided into true cholinesterase and false cholinesterase.True cholinesterase also claims acetylcholinesterase (acetylcholinesterase), is mainly present in cholinergic nerve endings synaptic cleft, assembles more in the wrinkle folding of particularly motor end plate postsynaptic membrane; Also be present in cholinergic neuron and in red corpuscle.This enzyme is the strongest for the Ach Pseudocholinesterase effect of physiological concentration, and specificity is also higher.Enzyme molecule hydrolyzable 3 × 10 molecule Ach.Pseudocholinesterase (PCHE) is present in serum or blood plasma, except acting on except vagusstoff, also can act on other cholines.When run into liver complex functionality decline, CHE(Pseudocholinesterase) and true cholinesterase concentration in blood decrease, its reduction degree and liver complex functionality degree of damage are proportionate.Choline fat is hydrolyzed to choline and organic acid by true cholinesterase, according to the number of its hydrolysate, can calculate CHE(Pseudocholinesterase clinically) activity.Therefore, Pseudocholinesterase is as follows at clinical application:
(1) hepatic diseases: Pseudocholinesterase is the index of reflection Hepatocyte anabolism function, in the hepatitis be in a bad way, its PCHE reduction is directly proportional to liver pathological changes degree, parallel with serum albumin; Then prognosis mala is pointed out as PCHE continues reduction when chronic hepatitis, liver cirrhosis, liver cancer; During hepatic insufficiency, PCHE obviously reduces.
(2) when heredity blood-serum P CHE is extremely sick, malnutritive, blood-serum P CHE all reduces.
(3) kidney disease (acatharsia), obesity, fatty liver, hyperthyroidism and heredity height PCHE mass formed by blood stasis person, blood-serum P CHE level all can raise.
(4) be the important means of assisting organophosphate poisoning diagnosis and prognostic to estimate.The pseudocholinesterase in red corpuscle in true cholinesterase and serum can be suppressed containing the sterilant of organophosphorus.During organophosphate poisoning, blood-serum P CHE reduces detection method:
Thiocholine+5,5 dithio-2-nitrobenzoic acid-----→ 5-sulfydryl-2-nitrobenzoic acid
Due to Butyryl thiocholine extremely unstable, be very easy to hydrolysis, have patent report (CN100557423C) substrate can stablize in an acidic solution, but in actual use procedure, effect be still undesirable, after long-time placement, substrate is still easy to hydrolysis.Cholinesterase assayautomated in the Cobas-Bio centrifugal analyzer.ClinicalChemistry August 1987 vol.33no.8 1460-1462 reports the method for stable Butyryl thiocholine; comprise inflated with nitrogen, with protective material etc.But these methods all thoroughly can not solve the problem of Butyryl thiocholine hydrolysis, substrate, after long-time placement, still progressively can be hydrolyzed, have a strong impact on blank and the sensitivity of reagent, thus finally affect detected result.
Summary of the invention
Technical problem to be solved by this invention is to overcome above-mentioned weak point, and research and design stablizes the material of substrate, improves reagent stability, for detecting the reagent of Pseudocholinesterase.
The invention provides a kind of Pseudocholinesterase detection kit, this test kit is by following reagent 1 He
Reagent 2, the ratio composition in 4 parts: 1 part:
Reagent 1:(1L) (reagent volume)
Reagent 2:(1L) (reagent volume)
Detection cholinesterase reagent box of the present invention, in use, first adds reagent 1 after detecting sample and adds reagent 2 again, can detect.
Another object of the present invention there is provided the preparation method of described cholinesterase reagent box: the method comprises the following steps:
(1) reagent 1:(1L is prepared) (reagent volume)
(1) in container, be first incorporated as the deionized water of total amount 80%;
(2) SODIUM PHOSPHATE, MONOBASIC, Sodium phosphate dibasic, hydroxy ethylene diphosphonic acid, sodium azide, 5 is added successively, 5 dithio-2-nitrobenzoic acids;
(3) deionized water to the cumulative volume finally adding residual content is that 1L mixes, and to obtain final product;
(2) reagent 2:(1L is prepared) (reagent volume)
In container, add tartrate, hydroxy ethylene diphosphonic acid, S-WAT, Sodium Benzoate, Butyryl thiocholine, ethylenediamine tetraacetic acid (EDTA), Virahol successively, it is that 1L mixes that deionized water adds to cumulative volume, to obtain final product.
The present inventor is confirmed by research trial: because the present invention adds Substrate Protection agent hydroxy ethylene diphosphonic acid and Virahol in test kit; overcoming prior art uses ethylenediamine tetraacetic acid (EDTA) to make the defect of protective material generation; very strong provide protection is had to the substrate in reagent; especially As time goes on, more obvious to the provide protection of substrate.Test kit Detection results of the present invention is good, easy to use, also can reduce medical expense, has larger clinical value.
Embodiment
Following examples agents useful for same raw material is all obtained by commercially available.
Embodiment 1 prepares Pseudocholinesterase detection kit
Composition: reagent 1:(1L) (reagent volume)
Reagent 2:(1L) (reagent volume)
Preparation:
(1) reagent 1:(1L is prepared) (reagent volume)
(1) in container, be first incorporated as the deionized water of total amount 80%;
(2) SODIUM PHOSPHATE, MONOBASIC, Sodium phosphate dibasic, hydroxy ethylene diphosphonic acid, sodium azide, 5 is added successively, 5 dithio-2-nitrobenzoic acids
(3) deionized water to the cumulative volume finally adding residual content is that 1L mixes, and to obtain final product;
(2) reagent 2:(1L is prepared) (reagent volume)
In container, add tartrate, hydroxy ethylene diphosphonic acid, S-WAT, Sodium Benzoate, Butyryl thiocholine, ethylenediamine tetraacetic acid (EDTA), Virahol successively, it is that 1L mixes that deionized water adds to cumulative volume, to obtain final product.
Embodiment 2
Composition: reagent 1:(1L) (reagent volume)
Reagent 2:(1L) (reagent volume)
Embodiment 3
Composition: reagent 1:(1L) (reagent volume)
Reagent 2:(1L) (reagent volume)
Preparation method is with embodiment 1.
Embodiment 4
Composition: reagent 1:(1L) (reagent volume)
Reagent 2:(1L) (reagent volume)
Preparation method is with embodiment 1.
Embodiment 5
The test kit Detection results test of the embodiment of the present invention 1:
Measuring method:
Step one: input parameter is in automatic clinical chemistry analyzer (Olympus 400): temperature 37 DEG C,
Predominant wavelength
405nm, commplementary wave length 520nm,
R1240ul(embodiment 1 reagent 1) .R2 60 ul(embodiment 1 reagent 2).
Detect sample 3ul(Chinese Shanghai Xinhua Hospital health check-up serum sample)
Reading point: 16-24
Step 2: before measuring, R1 and R2 is balanced to room temperature., then put into agent bin
Step 3 input scaling ratio: 7426 in use, reagent 1 adds reagent 2 after adding detection sample again, can detect.
Embodiment 6
Detect by the parameter of embodiment 5
Above-mentioned detected result shows that test kit of the present invention adds hydroxy ethylene diphosphonic acid and Virahol has very strong provide protection to the Butyryl thiocholine in reagent; As time goes on; can be more obvious to Protection of Substrates, the sample value adding protectant reagent after 12 months only reduces 5%.
The clinical correlation comparison test of embodiment 7 test kit of the present invention and DESAY's cholinesterase reagent box
The present embodiment 1 test kit and DESAY cholinesterase reagent box have been made clinical correlation and have been compared:
Get that the clinical sample of 30 routine different concns is parallel with commercially available DESAY's test kit (the reagent 1:4*64ml reagent 2:4*16ml) manufacturer (company of German DESAY) obtained to carry out detection and use instrument: Olympus 400
Measuring method
Step one: input parameter is in automatic biochemistry analyzer: temperature 37 DEG C, predominant wavelength
405nm, auxiliary wavelength 520nm,
Detect sample 2.5ul(Chinese Shanghai Xinhua Hospital health check-up serum sample)
Reading point: 24-27
In use, the present embodiment 1 kit reagent 1 adds reagent 2 after adding detection sample again, can detect.
Contrast agents: German DESAY cholinesterase reagent box (reagent 1200ul; Reagent 250ul)
Manufacturer (company of German DESAY);
Detect sample 2.5ul((Chinese Shanghai Xinhua Hospital health check-up serum sample)
Reading point: 24-27
Step 2: before measuring, R1 and R2 is balanced to room temperature., then put into agent bin
Step 3 input scaling ratio: 7426
Olympus 400 inputs above parameter, and in use, German DESAY cholinesterase reagent box reagent 1 adds reagent 2 after adding detection sample again, can detect.
Test-results:
According to clinical comparison data above, the clinical correlation of reagent of the present invention and DESAY's reagent is good, does not have difference.Further, reagent stability of the present invention is good, easy to use, may be used for clinical, also can reduce medical expense.
Claims (2)
1. Pseudocholinesterase detection kit, is characterized in that, described test kit, by following reagent 1 and reagent 2, forms in the ratio of 4 parts: 1 part:
Reagent 1:1L
Reagent 2:1L
2. prepare the method for Pseudocholinesterase detection kit as claimed in claim 1, it is characterized in that, the method comprises the following steps:
(1) reagent 1 is prepared:
Reagent 1:1L
(1) in container, be first incorporated as the deionized water of total amount 80%;
(2) SODIUM PHOSPHATE, MONOBASIC, Sodium phosphate dibasic, hydroxy ethylene diphosphonic acid, sodium azide, 5 is added successively, 5 dithio-2-nitrobenzoic acids;
(3) deionized water to the cumulative volume finally adding residual content is that 1L mixes, and to obtain final product; (2) reagent 2 is prepared:
Reagent 2:1L
In container, add tartrate, hydroxy ethylene diphosphonic acid, S-WAT, Sodium Benzoate, Butyryl thiocholine, Fucose, Virahol successively, it is that 1L mixes that deionized water adds to cumulative volume, to obtain final product.
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| Application Number | Priority Date | Filing Date | Title |
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| CN201210529456.8A CN103865978B (en) | 2012-12-10 | 2012-12-10 | Pseudocholinesterase detection kit and preparation method thereof |
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| Application Number | Priority Date | Filing Date | Title |
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| CN201210529456.8A CN103865978B (en) | 2012-12-10 | 2012-12-10 | Pseudocholinesterase detection kit and preparation method thereof |
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| CN103865978A CN103865978A (en) | 2014-06-18 |
| CN103865978B true CN103865978B (en) | 2015-09-16 |
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Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN106018308A (en) * | 2016-08-11 | 2016-10-12 | 山东博科生物产业有限公司 | Cholinesterase detection kit |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US3438866A (en) * | 1967-03-21 | 1969-04-15 | Pfizer & Co C | Serum cholinesterase test |
| CN1811393A (en) * | 2006-01-25 | 2006-08-02 | 长春迪瑞实业有限公司 | Liquid stablilized serum cholinesterase reagent |
| CN1978660A (en) * | 2005-12-08 | 2007-06-13 | 霍夫曼-拉罗奇有限公司 | Stabilized cholinesterase substrate solution |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP3051992B2 (en) * | 1990-11-29 | 2000-06-12 | 株式会社ヤトロン | Cholinesterase activity measurement reagent |
-
2012
- 2012-12-10 CN CN201210529456.8A patent/CN103865978B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US3438866A (en) * | 1967-03-21 | 1969-04-15 | Pfizer & Co C | Serum cholinesterase test |
| CN1978660A (en) * | 2005-12-08 | 2007-06-13 | 霍夫曼-拉罗奇有限公司 | Stabilized cholinesterase substrate solution |
| CN1811393A (en) * | 2006-01-25 | 2006-08-02 | 长春迪瑞实业有限公司 | Liquid stablilized serum cholinesterase reagent |
Non-Patent Citations (2)
| Title |
|---|
| New Findings about Ellman’s Method to Determine Cholinesterase Activity;Alena Komersova等;《Z. Naturforsch.》;20071231;第62卷;150-154 * |
| 血红蛋白存在下胆碱酯酶活性的5,5’一二硫双一2一硝基苯甲酸分光光度测定法;李前等;《中国卫生检验杂志》;20010630;第11卷(第3期);268-269 * |
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Address after: 200010 No. 2 Fuxing East Road, Shanghai, Huangpu District Patentee after: SHANGHAI FOSUN PHARMACEUTICAL(GROUP)CO., Ltd. Patentee after: Fosun diagnostic technology (Shanghai) Co., Ltd Address before: 200010 No. 2 Fuxing East Road, Shanghai, Huangpu District Patentee before: SHANGHAI FOSUN PHARMACEUTICAL(GROUP)CO., Ltd. Patentee before: Shanghai Fosun Changzheng Medical Science Co., Ltd |
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