CN102823579A - Blood protective agent, preparation method thereof and application - Google Patents

Blood protective agent, preparation method thereof and application Download PDF

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Publication number
CN102823579A
CN102823579A CN2012103068609A CN201210306860A CN102823579A CN 102823579 A CN102823579 A CN 102823579A CN 2012103068609 A CN2012103068609 A CN 2012103068609A CN 201210306860 A CN201210306860 A CN 201210306860A CN 102823579 A CN102823579 A CN 102823579A
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China
Prior art keywords
blood
hyclone
glycerine
solution
conservation agent
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Pending
Application number
CN2012103068609A
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Chinese (zh)
Inventor
陆维克
张蕾
吴淑江
陈金树
叶春生
王海剑
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Hangzhou Biotest Biotech Co Ltd
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Hangzhou Biotest Biotech Co Ltd
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Priority to CN2012103068609A priority Critical patent/CN102823579A/en
Publication of CN102823579A publication Critical patent/CN102823579A/en
Pending legal-status Critical Current

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Abstract

The invention discloses a blood protective agent and a preparation method thereof. The blood protective agent is a solution formed by dissolving a surface active agent, a preservative, fetal calf serum, glycerol, saccharides and sodium chloride in 0.605-60.5g/L of trihytdroxy methyl-aminomethane buffer solution, wherein the concentrations of the surface active agent, the preservative, the fetal calf serum, the glycerol, the saccharides and the sodium chloride in the solution are respectively 0.1-20gL, 0.01-2g/L, 10-200ml/L, 10-200ml/L, 5-50g/L and 9gL. According to the blood protective agent, after blood and the protective agent are uniformly mixed according to a certain proportion, an obtained mixture is placed at a temperature of 2-8 DEG C for preservation to ensure that serum and blood plasma can have a detection validity period longer than 120 days, so that the blood protective agent can be applied to the serum immunology and biological chemistry detection kits requiring to prepare various positive control, negative control and quantitative standard substances.

Description

A kind of blood conservation agent and preparation method thereof and application
Technical field
The invention belongs to biological technical field, relate in particular to a kind of blood conservation agent and preparation method thereof and application.
Background technology
The blood that in animal or human body, collects is called whole blood, whole blood is not added anticoagulant separate the liquid that obtains with blood cell and be called serum, and the liquid that in whole blood, adds behind the anticoagulant with the blood cell resulting separation is called blood plasma.Whole blood, serum, blood plasma all have certain pot-life; Under normal temperature liquid situation, be easy to lose efficacy; Therefore it is in demand needing the protectant of a kind of effective and feasible prolongation blood product pot-life, especially for preparation in various sero-immunities and the biochemistry detection kit or prepare positive control, negative control, critical value and various quantitative criterion article.
In blood or the blood plasma preservation process, prevent the method for blood coagulation at present, have following several kinds:
One, prevent that with chemicals blood coagulation from eliminating the effect of calcium ion with the calcium bond, make calcium in the blood change solubility into, but not Ionized chelate.The method is easy, is widely used.
(1) citric acid trisodium: two kinds of finished products that water of crystallization content is different are generally arranged: a kind of 2Na of being 3C 8H 5O 711H 2O, it such as becomes at concentration of oozing with blood plasma be 3.8%; A kind of in addition is Na 3C 8H 5O 72H 2O, the concentration of oozing such as becoming with blood plasma is 3.2% (2.5 ~ 4.0%), is alkalescence, and citric acid or its salt and calcium effect generate the chelate of solubility, and when processing 2.5% solution, PH is 7.5.Minimum anti-freezing concentration is 0.2%, and generally ultimate density should be 0.4 ~ 0.6% in blood, is the most general in the world for a long time anticoagulant.
(2) disodium citrate: wherein contain a large amount of citric acid ions, the PH of 3.5% solution is 4.5, compares with the citric acid trisodium, and its advantage is: simplified prescription and preparation formality thereof that liquid is preserved in configuration, the coking phenomenon of glucose when avoiding autoclave sterilization.
(3) three hydroxyl Sodium glutarates: can substitute sodium citrate, a little less than the anti-freezing power, the PH of its 7% solution is 4.3, and anti-freezing power is equivalent to 2.0 ~ 3.5% citrate solution, and toxicity is less, and Rollet is played booster action.
(4) EDTA-Na 2: the calcium complexing agent.It is a kind of very strong anticoagulant.1.5 gram EDTA-Na 2And the term of validity that 5% glucose solution is preserved 500ml blood at 4 degree is 28 days.
Two, preventing blood coagulation such as heparin with biologic product, is a kind of acid mucopolysaccharide, and its effect is the generation that stops fibrin ferment, thereby reaches the anti-freezing purpose.10mg (1000 unit) can make 100ml blood a couple of days not solidify, be mainly used in leucocyte and separate and preservation with hematoblastic, but because of its anti-freezing ability have the regular hour restriction with can not long preservation blood etc. shortcoming, its application is restricted.
Three, adopt the physical property method to prevent that blood coagulation Application of ion exchanger from removing the calcium ion in the blood, blood is not solidified.This blood does not contain anticoagulant substances, does not have the danger of poisoning, preserves but can not be used for blood.
Summary of the invention
The objective of the invention is to propose a kind of blood conservation agent; As solvent, wherein protectant comprises surfactant, preservative, hyclone to this protectant, glycerine, carbohydrate with water and TRIS buffer; Salt; After blood and protectant mixed with certain proportion, place 2 ~ 8 ℃ of preservations, the detection term of validity that makes serum, blood plasma have to reach more than 120 days.
The object of the invention can be realized through following technical scheme: a kind of blood conservation agent; Said protectant is dissolved in the solution that forms in the TRIS buffer of 0.605 ~ 60.5g/L by surfactant, preservative, hyclone, glycerine, carbohydrate, sodium chloride, wherein surfactant, preservative, hyclone, glycerine, carbohydrate, the concentration of sodium chloride in said solution be respectively 0.1 ~ 20gL, _ 0.01 ~ 2gL, 10 ~ 200ml/L, 10 ~ 200ml/L, 5 ~ 50gL, 9gL.
As preferably, surfactant of the present invention is Triton X-100 or C14-16 alkene sulfonic acid sodium, and preservative is ProClin 300 or Sodium azide, and carbohydrate is at least a in sucrose or trehalose or fructose or the dextran.
As preferably; Blood conservation agent of the present invention is to be dissolved in the solution that forms in the TRIS buffer of 6.05g/L by Triton X-100, ProClin 300, hyclone, glycerine, sucrose, sodium chloride, and Triton X-100, ProClin300, hyclone, glycerine, sucrose, the concentration of sodium chloride in said solution are respectively 1gL, 0.02g/L, 50ml/L, 20ml/L, 10g/L, 9g/L.
As preferably; Blood conservation agent of the present invention is to be dissolved in the solution that forms in the TRIS buffer of 1.21gL by Triton X-100, Sodium azide, hyclone, glycerine, trehalose, sodium chloride, and Triton X-100, Sodium azide, hyclone, glycerine, the concentration of trehalose in said solution are respectively 5gL, 0.02gL, 20ml/L, 50ml/L, 15g/L.
As preferably; Blood conservation agent of the present invention is to dissolve the solution that forms in the TRIS buffer of 2.42g/L in sight by Triton X-100, Sodium azide, hyclone, glycerine, dextran, sodium chloride, and Triton X-100, Sodium azide, hyclone, glycerine, the concentration of dextran in said solution are respectively 10gL, 0.02gL, 10ml/L, 100ml/L, 5g/L.
The method for preparing the blood conservation agent of the present invention, concrete steps are following:
1) takes by weighing each component by prescription;
2) distilled water that adds 700ml earlier is in container; Each component in the step 1) is added according to following order successively: surfactant, trishydroxymethylaminomethane, sodium chloride, preservative, hyclone, carbohydrate, glycerine; After mixing; Be settled to 1L, adjustment pH to 7.0 ~ 8.5 obtain the blood conservation agent.
The application of blood conservation agent of the present invention in stock's blood is preserved.
In the application of blood conservation agent according to the invention in stock's blood is preserved, typhoid fever, dengue fever, trypanosome blood preparation and the blood conservation agent of the gathering ratio with 1:1 ~ 1:2 is slowly fully mixed, place 2 ~ 8 ℃ of preservations.
The invention has the advantages that: 1. the pot-life that can prolong whole blood, serum, blood plasma reached more than 120 days; 2. be widely used, can be used for sero-immunity and biochemistry detection kit that various needs are equipped with positive control, negative control and various quantitative criterion article.
The practical implementation method
Following examples are used to explain the present invention, and are not used in restriction the present invention.
Instance 1
Typhoid fever IgM positive sample and 3 parts of negative blood preparations of typhoid fever with 3 parts of varying strengths make an experiment; Protectant consists of trishydroxymethylaminomethane (6.05g/L), Triton X-100 (1g/L), hyclone (50ml/L); Glycerine (20ml/L); Preservative ProClin300 (0.02g/L), sucrose (10g/L), sodium chloride (9g/L).Carry out check experiment at normal temperatures, wherein one group is blood preparation, and another group is for blood preparation and protectant mixed liquor (mixed proportion is 1:1), place 2 ~ 8 ℃ of preservation different times after, detect with a kind of typhoid fever detectable.
The positive detection result of specimen of table 1
Sample 0 day 14 days 28 days 56 days 90 days 120 days
Typhoid fever IgM positive sample 1 4+ 3+ 2+ Negative Negative Negative
Typhoid fever IgM positive sample 2 3+ 2+ 1+ Negative Negative Negative
Typhoid fever IgM positive sample 3 2+ 2+ 1+ Negative Negative Negative
Typhoid fever IgM positive sample 1+ protectant 4+ 4+ 4+ 4+ 3+ 3+
Typhoid fever IgM positive sample 2+ protectant 3+ 3+ 3+ 3+ 2+ 2+
Typhoid fever IgM positive sample 3+ protectant 2+ 2+ 2+ 2+ 1+ 1+
Annotate: "+" expression is positive, and numeral is big more, and the positive is strong more, is 5 to the maximum, and minimum is 1.
Can know by table 1, not add protectant positive sample and store 56 days at normal temperatures and all show negative findings, and add protectant all still positive findingses.
The negative detection result of specimen of table 2
Negative detection result of specimen
Sample 0 day 14 days 28 days 56 days 90 days 120 days
The negative sample 1 of typhoid fever IgM Negative Negative Negative Negative Negative Negative
The negative sample 2 of typhoid fever IgM Negative Negative Negative Negative Negative Negative
The negative sample 3 of typhoid fever IgM Negative Negative Negative Negative Negative Negative
The negative sample 1+ of typhoid fever IgM protectant Negative Negative Negative Negative Negative Negative
The negative sample 2+ of typhoid fever IgM protectant Negative Negative Negative Negative Negative Negative
The negative sample 3+ of typhoid fever IgM protectant Negative Negative Negative Negative Negative Negative
Can know that by table 2 do not add protectant negative sample and added protectant negative sample and all show negative findings, protectant can not cause false positive.
Instance 2
Dengue fever IgM positive sample with 3 parts of varying strengths makes an experiment; Protectant consists of trishydroxymethylaminomethane (1.21gL), Triton X-100 (5gL), hyclone (20ml/L); Glycerine (50ml/L); Preservative Sodium azide (0.02gL), trehalose (15g/L), sodium chloride (9g/L).Carry out check experiment, wherein one group is sample, and another group is for sample and protectant mixed liquor (mixed proportion is 1:2), place 2 ~ 8 ℃ of preservation different times after, detect with a kind of dengue fever detectable.
The positive detection result of specimen of table 3
Sample 0 day 15 days 30 days 2 months 4 months 6 months
Dengue fever IgM positive sample 1 3+ 2+ 1+ Negative Negative Negative
Dengue fever IgM positive sample 2 3+ 2+ 2+ 1+ Negative Negative
Dengue fever IgM positive sample 3 2+ 2+ 1+ Negative Negative Negative
Dengue fever IgM positive sample 1+ protectant 3+ 3+ 3+ 2+ 2+ 2+
Dengue fever IgM positive sample 2+ protectant 3+ 3+ 3+ 3+ 2+ 2+
Dengue fever IgM positive sample 3+ protectant 2+ 2+ 2+ 1+ 1+ 1+
Can know by table 3, not add protectant positive sample and store 4 monthly demonstration negative findingses at normal temperatures, and add 6 monthly still positive findingses of protectant preservation.
Instance 3
Make an experiment with 1 part of trypanosome IgG strong positive sample; Protectant consists of trishydroxymethylaminomethane (2.42g/L), Triton X-100 (10g/L), hyclone (10ml/L); Glycerine (100ml/L); Preservative Sodium azide (0.02gL), dextran (5g/L), sodium chloride (9gL).Carry out check experiment; Wherein one group is sample, and another group obtains mixed liquor, preservation at room temperature earlier in first day for sample and protectant with the mixed of 1:1; Preserved down at-20 ℃ again in second day; Beginning in the 3rd day is preserved at 40 ℃ of lower seals, and beginning in the 4th day detects with a kind of trypanosome detectable after preserving different time under 2 ~ 8 ℃.
The positive detection result of specimen of table 4
Sample 0 day 1 day 2 days 3 days 15 days 1 month 2 months 3 months
Trypanosome IgG positive sample 5+ 4+ 4+ 3+ 3+ ?2+ Negative Negative
Trypanosome IgG positive sample+protectant 5+ 5+ 5+ 5+ 4+ 4+ 4+ 3+
Can know that by table 4 do not add protectant positive sample after different temperatures change to store, tiring descends gradually, shows negative findings after 2 months, and add protectant preceding 3 days and tire constant, preserve 3 months still positive findingses afterwards.

Claims (8)

1. blood conservation agent; It is characterized in that; Said protectant is dissolved in the solution that forms in the TRIS buffer of 0.605 ~ 60.5gL by surfactant, preservative, hyclone, glycerine, carbohydrate, sodium chloride, and wherein surfactant, preservative, hyclone, glycerine, carbohydrate, the concentration of sodium chloride in said solution are respectively 0.1 ~ 20gL, 0.01 ~ 2gL, 10 ~ 200ml/L, 10 ~ 200ml/L, 5 ~ 50gL, 9g/L.
2. blood conservation agent according to claim 1; It is characterized in that; Described surfactant is Triton X-100 or C14-16 alkene sulfonic acid sodium, and preservative is ProClin 300 or Sodium azide, and carbohydrate is at least a in sucrose or trehalose or fructose or the dextran.
3. blood conservation agent according to claim 2; It is characterized in that; Described blood conservation agent is to be dissolved in the solution that forms in the TRIS buffer of 6.05g/L by Triton X-100, ProClin 300, hyclone, glycerine, sucrose, sodium chloride, and wherein Triton X-100, ProClin300, hyclone, glycerine, sucrose, the concentration of sodium chloride in said solution are respectively 1gL, 0.02gL, 50ml/L, 20ml/L, 10gL, 9g/L.
4. blood conservation agent according to claim 2; It is characterized in that; Described blood conservation agent is to be dissolved in the solution that forms in the TRIS buffer of 1.21g/L by Triton X-100, Sodium azide, hyclone, glycerine, trehalose, sodium chloride, and wherein Triton X-100, Sodium azide, hyclone, glycerine, the concentration of trehalose in said solution are respectively 5gL, 0.02g/L, 20ml/L, 50ml/L, 15gL.
5. blood conservation agent according to claim 2; It is characterized in that; Described blood conservation agent is to dissolve the solution that forms in the TRIS buffer of 2.42g/L in sight by Triton X-100, Sodium azide, hyclone, glycerine, dextran, sodium chloride, and wherein Triton X-100, Sodium azide, hyclone, glycerine, the concentration of dextran in said solution are respectively 10g/L, 0.02g/L, 10ml/L, 100ml/L, 5gL.
6. a method for preparing each said blood conservation agent of claim 1-5 is characterized in that, said preparation method's concrete steps are following:
1) takes by weighing each component by prescription;
2) distilled water that adds 700ml earlier is in container; Each component in the step 1) is added according to following order successively: surfactant, trishydroxymethylaminomethane, sodium chloride, preservative, hyclone, carbohydrate, glycerine; After mixing; Be settled to 1L, adjustment pH to 7.0 ~ 8.5 obtain the blood conservation agent.
7. the application of each described blood conservation agent of claim 1-5 in stock's blood is preserved.
8. application according to claim 7 is characterized in that, in said application, typhoid fever, dengue fever, trypanosome blood preparation and the blood conservation agent of the gathering ratio with 1:1 ~ 1:2 is slowly fully mixed, and places 2 ~ 8 ℃ of preservations.
CN2012103068609A 2012-08-27 2012-08-27 Blood protective agent, preparation method thereof and application Pending CN102823579A (en)

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Cited By (10)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103323308A (en) * 2013-05-23 2013-09-25 北京博晖创新光电技术股份有限公司 Reagent for processing trace whole blood and application thereof
CN104195221A (en) * 2014-08-18 2014-12-10 苏州康铭诚业医用科技有限公司 Composite stabilizer for glucose assay kit
CN104931317A (en) * 2015-06-11 2015-09-23 宁波美晶医疗技术有限公司 Blood pretreatment liquor
CN107083382A (en) * 2017-05-25 2017-08-22 广州奇辉生物科技有限公司 A kind of blood preseration agent and its application for being used to protect dissociative DNA
CN107302911A (en) * 2016-04-25 2017-10-31 中国农业科学院兰州兽医研究所 It is a kind of for preservation liquid of cattle and sheep esophagus-pharyngeal secretion and its production and use
CN107966555A (en) * 2017-11-23 2018-04-27 中山市创艺生化工程有限公司 It is a kind of to be used to stablize the storage liquid for preserving NAD or derivatives thereof
WO2018227359A1 (en) * 2017-06-13 2018-12-20 Chung Chin Sun A novel method for blood serum protein activity preservation
CN110352952A (en) * 2019-07-22 2019-10-22 苏州四正柏生物科技有限公司 A kind of composition saved for cell, aqueous compositions and its application
CN113243360A (en) * 2021-07-07 2021-08-13 天津德祥生物技术有限公司 Plasma preservation solution and application thereof
WO2022010916A1 (en) * 2020-07-07 2022-01-13 Truckee Applied Genomics, Llc Cell preservation reagents and methods of use

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CN101819202A (en) * 2010-05-06 2010-09-01 上海太阳生物技术有限公司 D-Dimer measuring kit (latex immunonephelometry method)
CN101970676A (en) * 2007-12-20 2011-02-09 雅培医护站股份有限公司 Compositions for forming immobiized biological layers for sensing
CN102628864A (en) * 2011-12-30 2012-08-08 北京九强生物技术股份有限公司 Kit for determining heart-type fatty acid binding protein in serum or urine by latex enhanced turbidimetric immunoassay

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CN101970676A (en) * 2007-12-20 2011-02-09 雅培医护站股份有限公司 Compositions for forming immobiized biological layers for sensing
CN101819202A (en) * 2010-05-06 2010-09-01 上海太阳生物技术有限公司 D-Dimer measuring kit (latex immunonephelometry method)
CN102628864A (en) * 2011-12-30 2012-08-08 北京九强生物技术股份有限公司 Kit for determining heart-type fatty acid binding protein in serum or urine by latex enhanced turbidimetric immunoassay

Cited By (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103323308A (en) * 2013-05-23 2013-09-25 北京博晖创新光电技术股份有限公司 Reagent for processing trace whole blood and application thereof
CN103323308B (en) * 2013-05-23 2016-01-06 北京博晖创新光电技术股份有限公司 A kind of reagent and application thereof processing micro whole blood
CN104195221A (en) * 2014-08-18 2014-12-10 苏州康铭诚业医用科技有限公司 Composite stabilizer for glucose assay kit
CN104931317A (en) * 2015-06-11 2015-09-23 宁波美晶医疗技术有限公司 Blood pretreatment liquor
CN107302911A (en) * 2016-04-25 2017-10-31 中国农业科学院兰州兽医研究所 It is a kind of for preservation liquid of cattle and sheep esophagus-pharyngeal secretion and its production and use
CN107083382A (en) * 2017-05-25 2017-08-22 广州奇辉生物科技有限公司 A kind of blood preseration agent and its application for being used to protect dissociative DNA
CN107083382B (en) * 2017-05-25 2018-07-31 广州奇辉生物科技有限公司 A kind of blood preseration agent and its application for protecting dissociative DNA
WO2018227359A1 (en) * 2017-06-13 2018-12-20 Chung Chin Sun A novel method for blood serum protein activity preservation
CN107966555A (en) * 2017-11-23 2018-04-27 中山市创艺生化工程有限公司 It is a kind of to be used to stablize the storage liquid for preserving NAD or derivatives thereof
CN110352952A (en) * 2019-07-22 2019-10-22 苏州四正柏生物科技有限公司 A kind of composition saved for cell, aqueous compositions and its application
WO2022010916A1 (en) * 2020-07-07 2022-01-13 Truckee Applied Genomics, Llc Cell preservation reagents and methods of use
CN113243360A (en) * 2021-07-07 2021-08-13 天津德祥生物技术有限公司 Plasma preservation solution and application thereof

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Application publication date: 20121219