CN104880350A - Preparation method for leucine aminopeptidase quality control product - Google Patents
Preparation method for leucine aminopeptidase quality control product Download PDFInfo
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- CN104880350A CN104880350A CN201510266759.9A CN201510266759A CN104880350A CN 104880350 A CN104880350 A CN 104880350A CN 201510266759 A CN201510266759 A CN 201510266759A CN 104880350 A CN104880350 A CN 104880350A
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Abstract
The invention discloses a preparation method for a leucine aminopeptidase quality control product. The preparation method comprises the following steps: freeze thawing a healthy person serum for 10-15 times, filtering and adjusting the pH value of filtrate to be 7.0-7.5; adding maleate and uniformly mixing; adding canavalin A and uniformly mixing, wherein in a final mixture, the concentration of maleate is 5-20 mmol/L and the concentration of canavalin A is 0.1-0.4 g/L; adding amino acid, polyethylene glycol, bovine serum albumin and a preservative in the mixed solution and uniformly mixing, wherein in a final mixture, the concentration of amino acid is 10-100 mmol/L, the concentration of polyethylene glycol is 1-10 g/L, the concentration of bovine serum albumin is 1-10 g/L and the concentration of the preservative is 0.2-0.5 g/L; sub-packaging and freeze-drying. The advantage of ensuring LAP (leucine aminopeptidase) determination stability is realized.
Description
Technical field
The present invention relates to technical field of medical examination, be specifically related to a kind of preparation method of leucine aminopeptidase quality-control product.
Background technology
Leucine aminopeptidase (Leucine aminopeptidase, LAP, EC 3.4.11.1) is a kind of protease, is distributed widely in the tissues such as liver, pancreas, kidney, when these tissues have pathology, all can occurs that serum LAP level raises.At present; measure the index that serum LAP is diagnosing hepatism; valuable to the diagnosis of the antidiastole of Cholestatic hepatitis, obstruction of biliary tract and cancer of pancreas, comparatively adenosine deaminase is high for the positive rate that detects of its liver cancer, sensitiveer than glutamyl transferase reflection liver diseases.The detection method of LAP is that LAP in sample is through the bright ammonia of catalyzing hydrolysis substrate L--P-nitroaniline, generate leucine and paranitroanilinum, reactive system rises in 405nm place absorbance, the speed risen by detecting it can draw the activity of LAP in sample, the LAP detection kit applying this method at home and abroad occurs all, but for want of international standard, cause testing result and may occur inconsistent phenomenon with manufacturer, reagent difference, be difficult to the stability ensureing that LAP measures, bring extreme difficulties to applying of this project.
In addition, in LAP Detection job management system, application quality-control product carries out Internal Quality Control and the quality assessment participated between room ensures laboratory detection result quality, reduces error, makes the result between different experiments room have the important measures of better comparability.So, provide a kind of and can ensure that the LAP quality-control product of LAP mensuration stability seems particularly important.
Summary of the invention
Technical matters to be solved by this invention is to provide a kind of preparation method that LAP measures the LAP quality-control product of stability that can ensure.
The technical solution adopted in the present invention is: a kind of preparation method of LAP quality-control product, comprises the following steps:
(1) Healthy Human Serum is adopted, multigelation (repeatedly freezing with thawing) 10-15 time, filtration, filtrate adjust ph to 7.0 ~ 7.5;
(2) in the serum (i.e. the filtrate of step 1) of above-mentioned adjusted pH value, add maleate, mix; Then add con A, mix; In the final mixture of gained, maleate concentration is 5 ~ 20mmol/L, the concentration of con A is 0.1 ~ 0.4g/L;
(3) in the mixed solution of above-mentioned steps (2), add amino acid, polyglycol, bovine serum albumin(BSA), antiseptic, mix; Amino acid/11 0 ~ 100mmol/L, polyglycol 1 ~ 10g/L, bovine serum albumin(BSA) 1-10g/L, antiseptic 0.2-0.5g/L in the final mixture of gained;
(4) packing, freeze-drying.
The preparation method of above-mentioned LAP quality-control product, the Healthy Human Serum described in step (1) can, from same individuality, also can be the pooled serum of Different Individual.
The preparation method of above-mentioned LAP quality-control product, the filtration described in step (1), is specially and uses fine and close filter paper, 0.45 μm of filter membrane and 0.22 μm of membrane filtration respectively (fine and close filter paper is exactly conventional filter paper, limits requirement without specification; Filtering through three roads makes product be in germ-free condition).
The preparation method of above-mentioned LAP quality-control product, the maleate described in step (2) is the one in maleic acid potassium or sodium maleate.
The preparation method of above-mentioned LAP quality-control product, the amino acid described in step (3) is the one of glycocoll, alanine and glutamic acid.
The preparation method of above-mentioned LAP quality-control product, the polyglycol described in step (3) is the one in polyglycol 3400, PEG 8000.
The preparation method of above-mentioned LAP quality-control product, the antiseptic described in step (3) is the one in Sodium azide or Proclin 300.
Advantage of the present invention and beneficial effect:
1, the present invention is because the Healthy Human Serum adopted, thus nature can avoid in patients serum with the interference of material.And Healthy Human Serum is material, source is sufficient, easily obtains, and can avoid contingent matrix effect to greatest extent, bacterial contamination, haemolysis, and fat is turbid waits nonspecific interference.
2, preparation technology is simple, and amount of additives is few, and production cost is low.
3, the present invention is by multigelation with regulate pH, effectively can remove lipoprotein, reduce differences between batches prepared by quality-control product, and add for subsequent species the pH value environment providing suitable; Add maleate and con A, in respective concentration range, protect the stability of the leucine aminopeptidase in serum, prevent it from degrading in subsequent treatment; The amino acid added, polyglycol, bovine serum albumin(BSA) antiseptic, being also the stability in order to protect the leucine aminopeptidase in serum in respective concentration range, preventing it from degrading in subsequent treatment.
Embodiment
Below by embodiment, the present invention is described in further detail, but the present invention is not only confined to following examples.
Embodiment 1
(1) the Healthy Human Serum 150ml eliminating infectious disease, common disease is collected, as sera stock, repeatedly to melt for interval is placed among room temperature and-20 DEG C of refrigerators respectively every 24 hours, freezing, after 20 days, careful separation goes out upper serum, first use fine and close Filter paper filtering 1 time, respectively filter 1 time with 0.45 μm of filter membrane and 0.22 μm of filter membrane respectively, regulate pH to 7.0;
(2) get the serum 100ml of above-mentioned adjusted pH value, add 10mmol/L maleic acid potassium, mix, then add 0.2g/L con A, mix;
(3) get above-mentioned (2) solution 100ml, add 20mmol/L glycocoll, 5g/L PEG 8000,5g/L bovine serum albumin(BSA), 0.3g/L Sodium azide, mixes;
(4) 1ml/ bottle packing, freeze-drying.
Embodiment 2
(1) the Healthy Human Serum 150ml eliminating infectious disease, common disease is collected, as sera stock, repeatedly to melt for interval is placed among room temperature and-20 DEG C of refrigerators respectively every 24 hours, freezing, after 30 days, careful separation goes out upper serum, first use fine and close Filter paper filtering 1 time, respectively filter 1 time with 0.45 μm of filter membrane and 0.22 μm of filter membrane respectively, regulate pH to 7.5;
(2) get the serum 100ml of above-mentioned adjusted pH value, add 20mmol/L sodium maleate, mix, then add 0.3g/L con A, mix;
(3) get above-mentioned (2) solution 100ml, add 50mmol/L glutamic acid, 8g/L polyglycol 3400,8g/L bovine serum albumin(BSA), 0.3g/L Sodium azide, mixes;
(4) 1ml/ bottle packing, freeze-drying.
It is as shown in table 1 below that quality-control product prepared by the above embodiment of the present invention preserves the result of 1 year in 2-8 DEG C of environment:
Table 1
| Month | Embodiment 1 (U/L) | Embodiment 2 (U/L) |
| 0 | 86.6 | 85.3 |
| 3 | 86.1 | 85.8 |
| 6 | 84.8 | 84.3 |
| 9 | 83.1 | 83.4 |
| 12 | 83.3 | 82.6 |
Detect data from above-mentioned table 1 can draw, illustrate that quality-control product of the present invention is stable and also can deposit the long period.
The raw material that embodiment is used, unless otherwise indicated, is commercially available industrial goods.
Above embodiment is to explanation of the present invention and explains further, instead of limitation of the present invention, and any amendment made within the scope of spirit of the present invention and right protection, all falls into protection scope of the present invention.
Claims (7)
1. a preparation method for leucine aminopeptidase quality-control product, is characterized in that: preparation process comprises:
(1) adopt Healthy Human Serum, multigelation 10-15 time, filter, filtrate adjust ph to 7.0 ~ 7.5;
(2) in the serum of above-mentioned adjusted pH value, add maleate, mix; Then add con A, mix; In the final mixture of gained, maleate concentration is 5 ~ 20mmol/L, the concentration of con A is 0.1 ~ 0.4g/L;
(3) in the mixed solution of above-mentioned steps (2), add amino acid, polyglycol, bovine serum albumin(BSA), antiseptic, mix; Amino acid/11 0 ~ 100mmol/L, polyglycol 1 ~ 10g/L, bovine serum albumin(BSA) 1-10g/L, antiseptic 0.2-0.5g/L in the final mixture of gained;
(4) packing, freeze-drying.
2. the preparation method of leucine aminopeptidase quality-control product according to claim 1, is characterized in that: the Healthy Human Serum described in step (1) is the pooled serum from same individuality or Different Individual.
3. the preparation method of leucine aminopeptidase quality-control product according to claim 1, is characterized in that: the filtration described in step (1), is specially and filters successively with fine and close filter paper, 0.45 μm of filter membrane and 0.22 μm of membrane filtration respectively.
4. the preparation method of leucine aminopeptidase quality-control product according to claim 1, is characterized in that: the maleate described in step (2) is the one in maleic acid potassium or sodium maleate.
5. the preparation method of leucine aminopeptidase quality-control product according to claim 1, is characterized in that: the amino acid described in step (3) is the one in glycocoll, alanine or glutamic acid.
6. the preparation method of leucine aminopeptidase quality-control product according to claim 1, is characterized in that: the polyglycol described in step (3) is the one in polyglycol 3400, PEG 8000.
7. the preparation method of leucine aminopeptidase quality-control product according to claim 1, is characterized in that: the antiseptic described in step (3) is the one in Sodium azide or Proclin 300.
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| CN106199020A (en) * | 2016-07-01 | 2016-12-07 | 长沙金域医学检验所有限公司 | A kind of method of manufacture and use thereof of hepatitis B surface antigen serum Internal Quality Control product |
| CN108802355A (en) * | 2018-06-15 | 2018-11-13 | 浙江省人民医院 | A kind of biochemistry detection quality-control product and quality control method |
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| CN106199020A (en) * | 2016-07-01 | 2016-12-07 | 长沙金域医学检验所有限公司 | A kind of method of manufacture and use thereof of hepatitis B surface antigen serum Internal Quality Control product |
| CN108802355A (en) * | 2018-06-15 | 2018-11-13 | 浙江省人民医院 | A kind of biochemistry detection quality-control product and quality control method |
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Address after: 315104 Ningbo, Yinzhou, Central District, No. Qiming South Road, No. 299 Applicant after: Meikang biological Polytron Technologies Inc Address before: 315104 Ningbo, Yinzhou, Central District, No. Qiming South Road, No. 299 Applicant before: Ningbo Meikang Biotechnology Co., Ltd. |
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Effective date of registration: 20190711 Address after: 311200 Floor 2, 873 Xinbei Road, Xiaoshan District, Hangzhou City, Zhejiang Province Patentee after: Hangzhou Mekongshengde Medical Laboratory Co., Ltd. Address before: 315104 No. 299 Qiming South Road, Yinzhou Central District, Ningbo City, Zhejiang Province Patentee before: Meikang biological Polytron Technologies Inc |