CN104498586A - Single reagent serum triglyceride detection reagent with strong stability - Google Patents
Single reagent serum triglyceride detection reagent with strong stability Download PDFInfo
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Abstract
The invention discloses a single reagent serum triglyceride detection reagent with strong stability, and belongs to the technical field of clinical in-vitro detection. The reagent adopts a 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES)-sodium glutamate buffer system, so that the system has a good buffer capacity; triethanolamine lauryl sulfate, a coenzyme FAD, polyvinyl alcohol AH-26 and other substances are added in the reagent and all effectively improve the stability of the reagent. Compared with a conventional single reagent triglyceride detection reagent, the reagent has more excellent stability and is more conducive to clinical popularization and use.
Description
Technical field
The present invention relates to the single reagent serum triglyceride detection reagent that a kind of stability is strong, belong to clinical vitro detection technical field.
Background technology
Triglyceride level (TG) is the chief component of blood fat, is the important energy supply material of body.Triglyceride level in food is at enteron aisle through steapsin digest and decompose, and after absorption, esterification is triglyceride level again, enters blood through lymphsystem.Liver is the main place of synthetic glycerine three ester, and fatty tissue is maximum triglyceride level storage vault.Under pathological conditions, the synthesis of various histocyte and the triglyceride level that preserves significantly increase.Content of triglyceride in detection by quantitative blood and histocyte is the conventional biochemical indicator of clinical, Clinical Basis, preclinical medicine biological study.
Current serum TG detection method is numerous, generally can be divided into the large class of chemical method, enzyme process and chromatography 3.Chemical method is with the TG in organic solvent extracting sample, after removing in extract the chaff interferences such as phosphatide, with basic hydrolysis (saponification) TG, generates formaldehyde with periodate oxidation glycerine, then surveys formaldehyde by color reaction.Methylene dichloride silicic acid off-color acid method (VanHande-Caslson method) more accurately, but operation more complicated, and technical requirements is high, is unsuitable for applying in routine clinical analysis; And chromatography is to the requirement height very of instrument and operator, and analysis cost is expensive, and therefore chemical method and chromatography are unfavorable for clinical expansion.
Domestic nearly all clinical detection mainly adopts enzyme process to detect TG content in serum, and its main operational principle is as follows: it is glycerine and lipid acid that lipase decomposes Triglycerides in Serum.Under ATP exists, Phosphoric acid glycerol estersization is generated glycerol 3-phosphate by glycerol kinase.The latter is generated phosphodihydroxyacetone and hydrogen peroxide by GPO oxidation.Hydrogen peroxide and peroxidase, 4-pyramidon carry out color reaction, generate coloured benzoquinone imine, are directly proportional to triglyceride concentration at 480 ~ 550 nm place absorbances.This method have easy fast, trace, advantage that precision is high, and high specificity, be easy to reach terminal, linearity range is wide, is conducive to clinical expansion.Generally many is on the market the TG detection reagent of double reagent, and the TG detection reagent of single reagent is due to its less stable, and reagent be extremely easily subject to environment pollution and not by market is accepted.
Summary of the invention
For solving problems of the prior art, the invention provides the single reagent serum levels of triglyceride detection reagent that a kind of stability is strong.
The present invention is by the following technical solutions:
The single reagent serum levels of triglyceride detection reagent that a kind of stability is strong, it is characterized in that, described reagent comprises: 50-60mmol/L 4-hydroxyethyl piperazine ethanesulfonic acid-Sodium Glutamate damping fluid (pH=7.4), 1-5g/L bovine serum albumin, 0.1-1g/L dodecyltriethanolamine sulfate, 0.1-1g/L peroxybenzoic acid sodium, 1-2g/L coenzyme F AD, 0.1-1g/L polyvinyl alcohol AH-26, 10-100mmol/L disodium ethylene diamine tetraacetate, 3.5-5mmol/L Sodium cholic acid, 17.5-20.5mo/Ll bitter salt, 1-3mmol/L 4-AA, 3.5-5mmol/L N, accelerine, 0.1-1g/L TritonX-100, 1-3KU/L lipoprotein lipase, 0.25-500KU/L glycerol kinase, 1-3KU/L peroxidase, 1-3KU/L glycerol-3-phosphate oxidase, 0.1-1g/L sodium azide.
Preferred as one of the present invention, described reagent comprises: 50mmol/L 4-hydroxyethyl piperazine ethanesulfonic acid-Sodium Glutamate damping fluid (pH=7.4), 3g/L bovine serum albumin, 0.5g/L dodecyltriethanolamine sulfate, 0.5g/L peroxybenzoic acid sodium, 1.5g/L coenzyme F AD, 0.5g/L polyvinyl alcohol AH-26, 50mmol/L disodium ethylene diamine tetraacetate, 4mmol/L Sodium cholic acid, 18mol/L bitter salt, 2mmol/L 4-AA, 4.5mmol/L N, accelerine, 0.5g/L TritonX-100, 2KU/L lipoprotein lipase, 300KU/L glycerol kinase, 2KU/L peroxidase, 2KU/L glycerol-3-phosphate oxidase, 0.5g/L sodium azide.
Each component is dissolved in 1L distilled water having prepared by the preparation method of reagent of the present invention successively.
The invention has the beneficial effects as follows:
1) the present invention have selected 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES)-Sodium Glutamate damping fluid, this buffer system is have good surge capability within the scope of 6.8-8.2 in pH value, make whole formula be in a kind of extremely stable state, improve the stability of reagent.
2) add dodecyltriethanolamine sulfate respectively in reagent of the present invention, it effectively can protect the stability of lipoprotein lipase; In addition, in reagent, add coenzyme F AD and polyvinyl alcohol AH-26, they can effectively in protected glycerol-3-phosphoric acid oxydase the active group of enzyme make enzyme not easy in inactivation, thus improve the stability of enzyme.
4) choose stability in the present invention comparatively strong, not labile DMA, as chromogen, substitutes original 4-chlorophenol, effectively slows down the disadvantage that reagent blank raises, thus ensure that the stability of reagent.
5) in reagent, also add the peroxybenzoic acid sodium with weak oxide character in the present invention, effectively can remove the interference of the reducing substances such as xitix, bilirubin to biochemical reagents reducing substances, thus improve immunity from interference and the stability of reagent.
Accompanying drawing explanation
Fig. 1 is embodiment of the present invention 1-3 and control group 7 days 37 DEG C of heat stability test result figure;
Fig. 2 is embodiment of the present invention 1-3 and control group 15 days reagent stability test result figure;
Fig. 3 is embodiment of the present invention 1-3 and control group 7 days 37 DEG C of thermostability reagent blank detected result figure;
Fig. 4 is embodiment of the present invention 1-3 and control group 15 days reagent stability reagent blank detected result figure.
Embodiment
Below in conjunction with specific embodiment, the present invention is described in further detail.
Embodiment 1
The single reagent serum levels of triglyceride detection reagent that a kind of stability is strong, described reagent comprises: 50mmol/L 4-hydroxyethyl piperazine ethanesulfonic acid-Sodium Glutamate damping fluid (pH=7.4), 1g/L bovine serum albumin, 0.1g/L dodecyltriethanolamine sulfate, 0.1g/L peroxybenzoic acid sodium, 1g/L coenzyme F AD, 0.1g/L polyvinyl alcohol AH-26, 10mmol/L disodium ethylene diamine tetraacetate, 3.5mmol/L Sodium cholic acid, 17.5mo/Ll bitter salt, 1mmol/L 4-AA, 3.5mmol/L N, accelerine, 0.1g/L TritonX-100, 1KU/L lipoprotein lipase, 0.25KU/L glycerol kinase, 1KU/L peroxidase, 1KU/L glycerol-3-phosphate oxidase, 0.1g/L sodium azide.
Above-mentioned each component is dissolved in 1L distilled water successively and has prepared.
Embodiment 2
The single reagent serum triglyceride detection reagent that a kind of stability is strong, described reagent comprises: 60mmol/L 4-hydroxyethyl piperazine ethanesulfonic acid-Sodium Glutamate damping fluid (pH=7.4), 5g/L bovine serum albumin, 1g/L dodecyltriethanolamine sulfate, 1g/L peroxybenzoic acid sodium, 2g/L coenzyme F AD, 1g/L polyvinyl alcohol AH-26, 100mmol/L disodium ethylene diamine tetraacetate, 5mmol/L Sodium cholic acid, 20.5mo/Ll bitter salt, 3mmol/L 4-AA, 5mmol/L N, accelerine, 1g/L TritonX-100, 3KU/L lipoprotein lipase, 500KU/L glycerol kinase, 3KU/L peroxidase, 3KU/L glycerol-3-phosphate oxidase, 1g/L sodium azide.
Above-mentioned each component is dissolved in 1L distilled water successively and has prepared.
Embodiment 3
The single reagent serum triglyceride detection reagent that stability is strong, described reagent comprises:
50mmol/L 4-hydroxyethyl piperazine ethanesulfonic acid-Sodium Glutamate damping fluid (pH=7.4), 3g/L bovine serum albumin, 0.5g/L dodecyltriethanolamine sulfate, 0.5g/L peroxybenzoic acid sodium, 1.5g/L coenzyme F AD, 0.5g/L polyvinyl alcohol AH-26, 50mmol/L disodium ethylene diamine tetraacetate, 4mmol/L Sodium cholic acid, 18mol/L bitter salt, 2mmol/L 4-AA, 4.5mmol/L N, accelerine, 0.5g/L TritonX-100, 2KU/L lipoprotein lipase, 300KU/L glycerol kinase, 2KU/L peroxidase, 2KU/L glycerol-3-phosphate oxidase, 0.5g/L sodium azide.
Said components is dissolved in 1L distilled water successively and has prepared.
Reagent obtained by embodiment of the present invention 1-3 is carried out stability test checking.Test is divided into uncork stability checking in 15 days and 7 days 37 DEG C of thermostability proof tests.
Test calibration object used and quality control product is respectively:
Calibration object: the content of Landau compound standard (848UN) wherein TG is 1.14mmol/L
Quality control product: Landau compound high level Quality Control: 620UE (TG target value: 2.86mmol/L, target value scope: 2.41-3.31mmol/L); Landau compound low value Quality Control: 897UN (TG target value: 1.09mmol/L, target value scope: 0.92-1.26mmol/L).
The concrete operation method of reagent stability proof test:
Using gained detection reagent in embodiment of the present invention 1-3 as test group, get the single reagent serum triglyceride detection reagent of a kind of certain company commercially available production as a control group, test group and control group reagent often organize two parts that ask for identical, portion does uncork stability test in 15 days, reagent is placed in 2-8 DEG C of refrigerator of instrument and (within 15 days, does not take out), as 15 days uncork Detection of Stability; Another part does 37 DEG C of heat stability test tests, closes and is placed on (every day only takes out when detection, and after detection, still sealing is put back in 37 DEG C of water-baths, continuous 7 days) in 37 DEG C of thermostat water baths, as 37 DEG C of thermostability checkings in 7 days.By reagent simultaneously on Toshiba-120 automatic clinical chemistry analyzer device, detect according to such as following table 1 method, and on instrument Criterion curve.Get the high and low value lyophilized powder quality control product of Landau respectively, after being uniformly dissolved, be divided into 15 parts separately ,-20 DEG C of storages, every day, high and low value Quality Control respectively got one, and tracing detection result, its tracking monitor trend is as Fig. 1 and Fig. 2.Detect the reagent blank of each group reagent every day, detected result is as Fig. 3, Fig. 4.
Table 1TG (triglyceride level) detection reagent detection method
Calculate TG(triglyceride level) content (μm ol/L)=(A mensuration/min ÷ A standard/min) × C standard.
No matter the TG detection reagent can found out in embodiment 1-3 by Fig. 1 and Fig. 2 is 7 days 37 DEG C of heat stability tests or reagent stability test in 15 days, and the stability of reagent is all good than contrast group reagent, is conducive to clinical expansion and uses.This mainly stablizes due to buffer system in reagent of the present invention, and add dodecyltriethanolamine sulfate respectively in reagent, it effectively can protect the stability of lipoprotein lipase; Simultaneously also add coenzyme F AD and polyvinyl alcohol AH-26 in reagent, they can effectively in protected glycerol-3-phosphoric acid oxydase the active group of enzyme make enzyme not easy in inactivation, thus improve the stability of enzyme.In addition, choose stability in the present invention comparatively strong, not labile DMA, as chromogen, substitutes original 4-chlorophenol, effectively slows down the disadvantage that reagent blank raises, thus ensure that the stability of reagent.
Can be found by Fig. 3 and Fig. 4, control group reagent attenuation ratio is very fast, and reagent blank is very easy to raise, the non-constant of stability of reagent self, and reagent blank absorbancy is when being greater than 0.08, and reagent just cannot use.And embodiment of the present invention 1-3 gained reagent, its 7 days 37 DEG C of thermostabilitys and 15 days reagent stability, detected result is all very stable, there is not the situation decayed in reagent, although reagent blank also has rising to a certain degree, ascensional range is little, not more than 0.08, reagent is still highly stable, is very beneficial for clinical expansion and uses.
Claims (2)
1. the single reagent serum levels of triglyceride detection reagent that a stability is strong, it is characterized in that, described reagent comprises: 50-60mmol/L 4-hydroxyethyl piperazine ethanesulfonic acid-Sodium Glutamate damping fluid (pH=7.4), 1-5g/L bovine serum albumin, 0.1-1g/L dodecyltriethanolamine sulfate, 0.1-1g/L peroxybenzoic acid sodium, 1-2g/L coenzyme F AD, 0.1-1g/L polyvinyl alcohol AH-26, 10-100mmol/L disodium ethylene diamine tetraacetate, 3.5-5mmol/L Sodium cholic acid, 17.5-20.5mo/Ll bitter salt, 1-3mmol/L 4-AA, 3.5-5mmol/L N, accelerine, 0.1-1g/L TritonX-100, 1-3KU/L lipoprotein lipase, 0.25-500KU/L glycerol kinase, 1-3KU/L peroxidase, 1-3KU/L glycerol-3-phosphate oxidase, 0.1-1g/L sodium azide.
2. single reagent serum triglyceride detection reagent according to claim 1, it is characterized in that, described reagent comprises: 50mmol/L 4-hydroxyethyl piperazine ethanesulfonic acid-Sodium Glutamate damping fluid (pH=7.4), 3g/L bovine serum albumin, 0.5g/L dodecyltriethanolamine sulfate, 0.5g/L peroxybenzoic acid sodium, 1.5g/L coenzyme F AD, 0.5g/L polyvinyl alcohol AH-26, 50mmol/L disodium ethylene diamine tetraacetate, 4mmol/L Sodium cholic acid, 18mol/L bitter salt, 2mmol/L 4-AA, 4.5mmol/L N, accelerine, 0.5g/L TritonX-100, 2KU/L lipoprotein lipase, 300KU/L glycerol kinase, 2KU/L peroxidase, 2KU/L glycerol-3-phosphate oxidase, 0.5g/L sodium azide.
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107884401A (en) * | 2017-11-13 | 2018-04-06 | 天津市宝坻区人民医院 | Eliminate the glucose oxidase assay method of piarhemia interference |
| CN108467882A (en) * | 2018-03-30 | 2018-08-31 | 潍坊市康华生物技术有限公司 | A kind of triglyceride detection kit |
| CN108949903A (en) * | 2017-05-17 | 2018-12-07 | 广州市伊川生物科技有限公司 | A kind of triglyceride determination kit and its measuring method |
| CN110938607A (en) * | 2019-12-18 | 2020-03-31 | 美康生物科技股份有限公司 | Glycerol-3-phosphate oxidase with good thermal stability and application thereof in kit |
| CN116718785A (en) * | 2023-06-28 | 2023-09-08 | 河南健士莱杰医疗科技有限公司 | Triglyceride kit with high stability and preparation method thereof |
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| CN102706818A (en) * | 2012-06-05 | 2012-10-03 | 宁波美康生物科技股份有限公司 | Enzymatic triglyceride measuring method and measuring reagent |
| CN104165992A (en) * | 2014-07-28 | 2014-11-26 | 武汉璟泓万方堂医药科技股份有限公司 | Kit for detecting apolipoprotein A5 for diagnosis of hypertriglyceridemia |
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| CN1446925A (en) * | 2003-03-24 | 2003-10-08 | 肖洪武 | Single stable colorimetric reagent of enzyme in liquid state and its application |
| CN102706818A (en) * | 2012-06-05 | 2012-10-03 | 宁波美康生物科技股份有限公司 | Enzymatic triglyceride measuring method and measuring reagent |
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108949903A (en) * | 2017-05-17 | 2018-12-07 | 广州市伊川生物科技有限公司 | A kind of triglyceride determination kit and its measuring method |
| CN107884401A (en) * | 2017-11-13 | 2018-04-06 | 天津市宝坻区人民医院 | Eliminate the glucose oxidase assay method of piarhemia interference |
| CN107884401B (en) * | 2017-11-13 | 2020-03-24 | 天津市宝坻区人民医院 | Glucose oxidase determination method for eliminating lipemia interference |
| CN108467882A (en) * | 2018-03-30 | 2018-08-31 | 潍坊市康华生物技术有限公司 | A kind of triglyceride detection kit |
| CN110938607A (en) * | 2019-12-18 | 2020-03-31 | 美康生物科技股份有限公司 | Glycerol-3-phosphate oxidase with good thermal stability and application thereof in kit |
| CN110938607B (en) * | 2019-12-18 | 2023-03-28 | 美康生物科技股份有限公司 | Glycerol-3-phosphate oxidase with good thermal stability and application thereof in kit |
| CN116718785A (en) * | 2023-06-28 | 2023-09-08 | 河南健士莱杰医疗科技有限公司 | Triglyceride kit with high stability and preparation method thereof |
| CN116718785B (en) * | 2023-06-28 | 2024-07-05 | 河南杰瑞生物工程有限公司 | Triglyceride kit with high stability and preparation method thereof |
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