CN118150844A - Liquid quality control product of glycosylated hemoglobin and preparation method thereof - Google Patents
Liquid quality control product of glycosylated hemoglobin and preparation method thereof Download PDFInfo
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- CN118150844A CN118150844A CN202410277231.0A CN202410277231A CN118150844A CN 118150844 A CN118150844 A CN 118150844A CN 202410277231 A CN202410277231 A CN 202410277231A CN 118150844 A CN118150844 A CN 118150844A
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- 102000017011 Glycated Hemoglobin A Human genes 0.000 title claims abstract description 53
- 108010014663 Glycated Hemoglobin A Proteins 0.000 title claims abstract description 53
- 238000003908 quality control method Methods 0.000 title claims abstract description 46
- 239000007788 liquid Substances 0.000 title claims abstract description 41
- 238000002360 preparation method Methods 0.000 title claims abstract description 17
- 239000000047 product Substances 0.000 claims abstract description 44
- 210000003743 erythrocyte Anatomy 0.000 claims abstract description 36
- 238000005406 washing Methods 0.000 claims abstract description 20
- 239000007853 buffer solution Substances 0.000 claims abstract description 19
- 239000003761 preservation solution Substances 0.000 claims abstract description 19
- 239000006228 supernatant Substances 0.000 claims abstract description 14
- 210000004369 blood Anatomy 0.000 claims abstract description 10
- 239000008280 blood Substances 0.000 claims abstract description 10
- 238000004321 preservation Methods 0.000 claims abstract description 8
- 238000005119 centrifugation Methods 0.000 claims abstract description 7
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 24
- 239000011780 sodium chloride Substances 0.000 claims description 12
- 238000000034 method Methods 0.000 claims description 9
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims description 7
- 239000008103 glucose Substances 0.000 claims description 7
- YASYEJJMZJALEJ-UHFFFAOYSA-N Citric acid monohydrate Chemical compound O.OC(=O)CC(O)(C(O)=O)CC(O)=O YASYEJJMZJALEJ-UHFFFAOYSA-N 0.000 claims description 6
- 229910019142 PO4 Inorganic materials 0.000 claims description 6
- 229960002303 citric acid monohydrate Drugs 0.000 claims description 6
- 239000000203 mixture Substances 0.000 claims description 6
- 239000008055 phosphate buffer solution Substances 0.000 claims description 6
- 239000011734 sodium Substances 0.000 claims description 6
- NLJMYIDDQXHKNR-UHFFFAOYSA-K sodium citrate Chemical compound O.O.[Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NLJMYIDDQXHKNR-UHFFFAOYSA-K 0.000 claims description 6
- 229960000999 sodium citrate dihydrate Drugs 0.000 claims description 6
- 239000000463 material Substances 0.000 claims 1
- 238000001514 detection method Methods 0.000 abstract description 8
- 238000004519 manufacturing process Methods 0.000 abstract description 6
- 238000003860 storage Methods 0.000 abstract description 6
- 230000007547 defect Effects 0.000 abstract description 5
- 239000000523 sample Substances 0.000 description 8
- 238000005259 measurement Methods 0.000 description 6
- 238000012360 testing method Methods 0.000 description 5
- 239000000843 powder Substances 0.000 description 3
- 230000002354 daily effect Effects 0.000 description 2
- 206010012601 diabetes mellitus Diseases 0.000 description 2
- 238000004108 freeze drying Methods 0.000 description 2
- 230000013595 glycosylation Effects 0.000 description 2
- 238000006206 glycosylation reaction Methods 0.000 description 2
- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Chemical compound N1C(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(NC(=O)CN)C(C)CC)CSSCC(C(NC(CO)C(=O)NC(CC(C)C)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CCC(N)=O)C(=O)NC(CC(C)C)C(=O)NC(CCC(O)=O)C(=O)NC(CC(N)=O)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CSSCC(NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2C=CC(O)=CC=2)NC(=O)C(CC(C)C)NC(=O)C(C)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2NC=NC=2)NC(=O)C(CO)NC(=O)CNC2=O)C(=O)NCC(=O)NC(CCC(O)=O)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC(O)=CC=3)C(=O)NC(C(C)O)C(=O)N3C(CCC3)C(=O)NC(CCCCN)C(=O)NC(C)C(O)=O)C(=O)NC(CC(N)=O)C(O)=O)=O)NC(=O)C(C(C)CC)NC(=O)C(CO)NC(=O)C(C(C)O)NC(=O)C1CSSCC2NC(=O)C(CC(C)C)NC(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(N)CC=1C=CC=CC=1)C(C)C)CC1=CN=CN1 NOESYZHRGYRDHS-UHFFFAOYSA-N 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- 102000001554 Hemoglobins Human genes 0.000 description 1
- 108010054147 Hemoglobins Proteins 0.000 description 1
- 102000004877 Insulin Human genes 0.000 description 1
- 108090001061 Insulin Proteins 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000003203 everyday effect Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000003317 immunochromatography Methods 0.000 description 1
- 229940125396 insulin Drugs 0.000 description 1
- 238000004255 ion exchange chromatography Methods 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 239000013610 patient sample Substances 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 238000000275 quality assurance Methods 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
Abstract
The invention discloses a liquid quality control product of glycosylated hemoglobin and a preparation method thereof, comprising the following steps: 1) Taking a human blood sample for centrifugation, and discarding the supernatant to obtain lower-layer red blood cells; 2) Adding buffer solution for washing, centrifuging, repeatedly washing, and discarding supernatant to obtain lower-layer washed red blood cells; 3) Adding a preservation solution into the red blood cells obtained in the step 2), storing for 10-14 hours at the temperature of-70 ℃ to-80 ℃, and then transferring to 2-8 ℃ for preservation to obtain a liquid quality control product of glycosylated hemoglobin; the liquid quality control product of the glycosylated hemoglobin overcomes the defect that the glycosylated hemoglobin needs to be freeze-dried for storage, improves the production and storage conditions of the quality control product, simplifies the preparation steps of the liquid quality control product of the glycosylated hemoglobin, and provides convenience for project detection in the medical field.
Description
Technical Field
The invention relates to the technical field of immunochromatography detection, in particular to a liquid quality control product of glycosylated hemoglobin and a preparation method thereof.
Background
Glycosylated hemoglobin (GHb) is the product of glycosylation of hemoglobin. The glycosylated hemoglobin can reflect the blood glucose level within 120 days before the detection stably and reliably, and is not influenced by factors such as blood drawing time, fasting, insulin use and the like. Glycosylated hemoglobin (GHb) is composed of HbA1a, hbA1b, hbA1c, wherein HbA1c occupies about 70%, and is structurally stable, and thus is used as a monitor index for diabetes control. Currently, glycosylated hemoglobin is widely used in clinic as an effective index for screening, diagnosing, controlling blood sugar and evaluating curative effect of diabetes.
Indoor quality control is an important component in laboratory quality assurance systems, the purpose of which is to ensure the stability and reliability of the measurement results for each patient sample. Firstly, the precision is high, namely the repeatability of the measurement result is good, the change of the measurement result in a laboratory every day is small, and the influence caused by random errors is eliminated or reduced; and secondly, the accuracy is high, namely the measurement result is correct and approaches to the true value. Mainly eliminating or reducing the influence of system errors. The quality control product plays an important role in the quality control work of clinical immune rooms. Accurate measurement of glycosylated hemoglobin requires a stable quality control to monitor the quality of the instrument and the reagent detection system. Because of the defects of easy oxidation and further glycosylation of glycosylated hemoglobin, most of glycosylated hemoglobin control products commonly used in the market are freeze-dried products, and a freeze-drying process is needed, so that the production conditions are harsh. Although the freeze-dried powder has the advantages of long storage time under normal temperature, convenient transportation and the like, the freeze-dried powder has the defects of quick use after reconstitution, accurate addition of quantitative distilled water, constant turbidity and high price of the reconstituted quality control product.
The existing preparation method of the liquid quality control product of the glycosylated hemoglobin is complicated, and is suitable for mass production of the glycosylated hemoglobin control product with a specific HbA1c value, and for a general laboratory, the problem is considered how to conveniently and rapidly obtain the liquid quality control product of the glycosylated hemoglobin, and the liquid quality control product of the glycosylated hemoglobin for daily quality control of the laboratory can be obtained without adopting steps such as ion chromatography or adding layering liquid into erythrocytes for layering.
Disclosure of Invention
The invention aims to overcome the defects of the prior art and provides a liquid quality control product of glycosylated hemoglobin and a preparation method thereof.
The aim of the invention is achieved by the following technical scheme: a preparation method of a liquid quality control product of glycosylated hemoglobin comprises the following steps:
1) Taking a human blood sample for centrifugation, and discarding the supernatant to obtain lower-layer red blood cells;
2) Adding buffer solution for washing, centrifuging, repeatedly washing, and discarding supernatant to obtain lower-layer washed red blood cells;
3) Adding a preservation solution into the red blood cells obtained in the step 2), storing for 10-14 hours at the temperature of-70 ℃ to-80 ℃, and then transferring to 2-8 ℃ for preservation to obtain a liquid quality control product of glycosylated hemoglobin;
In the step 1), the centrifugal speed is 2000 rpm-4000 rpm/min, and the centrifugal time is 5-20 min;
In the step 2), the buffer solution is phosphate buffer solution, and the specific composition is as follows: 5.0-8.0 g/L NaCl, 2.5-4.5 g/L Na 2HPO4•12H2 O, 0.4-0.8 g/L anhydrous KH 2PO4;
The volume ratio of the lower layer red blood cells to the buffer solution in the step 2) is 1:8-12;
In the step 2), the rotating speed of the centrifugal machine is the same as that of the centrifugal machine in the step 1), and the centrifugal washing is carried out for 2-5 times in the step 2);
In the step 3), the preservation solution comprises the following specific components: 5.0-10.0 g/L sodium citrate dihydrate, 0.4-0.8 g/L citric acid monohydrate, 16-22 g/L glucose, 5.0-8.0 g/L NaCl;
the invention has the following advantages:
1. The invention provides a liquid quality control product of glycosylated hemoglobin, which overcomes the defect that the glycosylated hemoglobin control product needs freeze-drying storage, improves the production and storage conditions of the quality control product, simplifies the preparation steps of the liquid quality control product of glycosylated hemoglobin, and provides convenience for project detection in the medical field.
2. The liquid glycosylated hemoglobin control provided by the invention can keep the stability of the protein content in a liquid state, and the protein content of the liquid glycosylated hemoglobin control can be kept unchanged within 6 months under the condition of 2-8 ℃ through test measurement, and the protein content of the liquid glycosylated hemoglobin control is kept unchanged under the condition of normal temperature for 7 days; compared with the quality control product of the freeze-dried powder sold in the market, the quality control product of the invention has the advantages of good stability, simple production condition, low cost, no need of cold chain transportation and great reduction of production, transportation and storage costs.
3. The preparation method of the liquid glycosylated hemoglobin control provided by the invention is simple and quick, can meet the daily quality control requirement of a laboratory, reduces the experiment cost and shortens the detection time; in the preparation method, a preservation solution is added into the washed red blood cells, the red blood cells are directly preserved at the temperature of 2 ℃ to 8 ℃ and frozen for 10 hours to 12 hours at the temperature of 70 ℃ to 80 ℃ below zero, and then the red blood cells are transferred to the temperature of 2 ℃ to 8 ℃ for preservation, so that the stability of the detection result of the glycosylated hemoglobin is different, and the former can be only stabilized for a short time.
Detailed Description
For the purpose of making the objects, technical solutions and advantages of the embodiments of the present invention more apparent, the technical solutions of the embodiments of the present invention will be clearly and completely described below, and it is apparent that the described embodiments are some embodiments of the present invention, but not all embodiments.
Thus, the following detailed description of the embodiments of the invention is not intended to limit the scope of the invention, as claimed, but is merely representative of selected embodiments of the invention. All other embodiments, based on the embodiments of the invention, which are apparent to those of ordinary skill in the art without inventive faculty, are intended to be within the scope of the invention.
In addition, the embodiments of the present invention and the features of the embodiments may be combined with each other without collision.
Example 1: a preparation method of a liquid quality control product of glycosylated hemoglobin comprises the following steps:
1) Taking a human blood sample for centrifugation, and discarding the supernatant to obtain lower-layer red blood cells;
2) Adding buffer solution for washing, centrifuging, repeatedly washing, and discarding supernatant to obtain lower-layer washed red blood cells;
3) Adding a preservation solution into the red blood cells obtained in the step 2), storing for 10-14 hours at the temperature of-70 ℃ to-80 ℃, and then transferring to 2-8 ℃ for preservation to obtain a liquid quality control product of glycosylated hemoglobin;
In the step 1), the centrifugal speed is 2000 rpm-4000 rpm/min, and the centrifugal time is 5-20 min;
In the step 2), the buffer solution is phosphate buffer solution, and the specific composition is as follows: 5.2g/L NaCl, 3.4g/L Na 2HPO4•12H2 O, 0.52g/L anhydrous KH 2PO4;
The volume ratio of the lower layer red blood cells to the buffer solution in the step 2) is 1:9;
In the step 2), the rotating speed of the centrifugal machine is the same as that of the centrifugal machine in the step 1), and the centrifugal washing is carried out for 2-5 times in the step 2);
in the step 3), the preservation solution comprises the following specific components: 7.5g/L sodium citrate dihydrate, 0.45g/L citric acid monohydrate, 21.2g/L glucose, 6.3g/L NaCl;
In the step 3), the volume ratio of the red blood cells to the preservation solution is 1:5.
Example 2: a preparation method of a liquid quality control product of glycosylated hemoglobin comprises the following steps:
1) Taking a human blood sample for centrifugation, and discarding the supernatant to obtain lower-layer red blood cells;
2) Adding buffer solution for washing, centrifuging, repeatedly washing, and discarding supernatant to obtain lower-layer washed red blood cells;
3) Adding a preservation solution into the red blood cells obtained in the step 2), storing for 14 hours at the temperature of-70 ℃, and then transferring to the temperature of 2 ℃ for preservation to obtain a liquid quality control product of glycosylated hemoglobin;
in the step 1), the centrifugal speed is 2000 rpm/min, and the centrifugal time is 20min;
in the step 2), the buffer solution is phosphate buffer solution, and the specific composition is as follows: 6.5g/L NaCl, 3.0g/L Na 2HPO4•12H2 O, 0.45g/L anhydrous KH 2PO4;
The volume ratio of the lower layer red blood cells to the buffer solution in the step 2) is 1:8;
in the step 2), the rotation speed of the centrifugal machine is the same as that of the centrifugal machine in the step 1), and the centrifugal washing is carried out for 2 times in the step 2);
in the step 3), the preservation solution comprises the following specific components: 7.0g/L sodium citrate dihydrate, 0.5g/L citric acid monohydrate, 19.5g/L glucose, 3.4g/L NaCl;
in the step 3), the volume ratio of the red blood cells to the preservation solution is 1:10.
Example 3: a preparation method of a liquid quality control product of glycosylated hemoglobin comprises the following steps:
1) Taking a human blood sample for centrifugation, and discarding the supernatant to obtain lower-layer red blood cells;
2) Adding buffer solution for washing, centrifuging, repeatedly washing, and discarding supernatant to obtain lower-layer washed red blood cells;
3) Adding a preservation solution into the red blood cells obtained in the step 2), storing for 10 hours at the temperature of minus 80 ℃, and then transferring to 8 ℃ for preservation to obtain a liquid quality control product of glycosylated hemoglobin;
In the step 1), the centrifugal speed is 4000rpm/min, and the centrifugal time is 5min;
In the step 2), the buffer solution is phosphate buffer solution, and the specific composition is as follows: 6.5g/L NaCl, 3.65g/L Na 2HPO4•12H2 O, 0.55g/L anhydrous KH 2PO4;
the volume ratio of the lower layer red blood cells to the buffer solution in the step 2) is 3:1;
in the step 2), the rotating speed of the centrifugal machine is the same as that of the centrifugal machine in the step 1), and the centrifugal washing is carried out for 5 times in the step 2);
In the step 3), the preservation solution comprises the following specific components: 6.5g/L sodium citrate dihydrate, 0.45g/L citric acid monohydrate, 18.6g/L glucose, 4.2g/L NaCl;
In the step 3), the volume ratio of the red blood cells to the preservation solution is 10:1.
Example 4: a preparation method of a liquid quality control product of glycosylated hemoglobin comprises the following steps:
1) Taking a human blood sample for centrifugation, and discarding the supernatant to obtain lower-layer red blood cells;
2) Adding buffer solution for washing, centrifuging, repeatedly washing, and discarding supernatant to obtain lower-layer washed red blood cells;
3) Adding a preservation solution into the red blood cells obtained in the step 2), storing for 12 hours at the temperature of 75 ℃ below zero, and then transferring to 5 ℃ for preservation to obtain a liquid quality control product of glycosylated hemoglobin;
In the step 1), the centrifugal rotating speed is 34000rpm/min, and the centrifugal time is 12min;
In the step 2), the buffer solution is phosphate buffer solution, and the specific composition is as follows: 6.0g/L NaCl, 4.2g/L Na 2HPO4•12H2 O, 0.52g/L anhydrous KH 2PO4;
the volume ratio of the lower layer red blood cells to the buffer solution in the step 2) is 1:10;
In the step 2), the rotation speed of the centrifugal machine is the same as that of the centrifugal machine in the step 1), and the centrifugal washing is carried out for 4 times in the step 2);
In the step 3), the preservation solution comprises the following specific components: 7.5g/L sodium citrate dihydrate, 0.42g/L citric acid monohydrate, 19.6g/L glucose, 4.2g/L NaCl;
In the step 3), the volume ratio of the red blood cells to the preservation solution is 1:5.
Example 4: a liquid quality control product of glycosylated hemoglobin is prepared by the method of example 4, and protein content test values of samples with different initial values stored for 1-6 months are shown in the following tables 1-3.
Table 1 sample test value with initial value of 5.5%
Table 2 sample test value with initial value of 7.82%
Table 3 sample test value with initial value of 9.36%
From the results shown in tables 1 to 3, it can be seen that the liquid quality control product of glycosylated hemoglobin of this example can maintain the protein content unchanged within 6 months, and is suitable for rapid laboratory preparation and for glycosylated hemoglobin detection.
Although the present invention has been described with reference to the foregoing embodiments, it will be apparent to those skilled in the art that modifications may be made to the embodiments described, or equivalents may be substituted for elements thereof, and any modifications, equivalents, improvements and changes may be made without departing from the spirit and principles of the present invention.
Claims (8)
1. A preparation method of a liquid quality control product of glycosylated hemoglobin is characterized by comprising the following steps: the method comprises the following steps:
1) Taking a human blood sample for centrifugation, and discarding the supernatant to obtain lower-layer red blood cells;
2) Adding buffer solution for washing, centrifuging, repeatedly washing, and discarding supernatant to obtain lower-layer washed red blood cells;
3) Adding preservation solution into the red blood cells obtained in the step 2), storing for 10-14 hours at the temperature of-70 ℃ to-80 ℃, and then transferring to 2-8 ℃ for preservation, thus obtaining the liquid quality control product of glycosylated hemoglobin.
2. The method for preparing a liquid quality control product of glycosylated hemoglobin according to claim 1, wherein: in the step 1), the centrifugal speed is 2000 rpm-4000 rpm/min, and the centrifugal time is 5-20 min.
3. The method for preparing a liquid quality control product of glycosylated hemoglobin according to claim 1, wherein:
In the step 2), the buffer solution is phosphate buffer solution, and the specific composition is as follows: 5.0-8.0 g/L NaCl, 2.5-4.5 g/L Na 2HPO4•12H2 O, 0.4-0.8 g/L anhydrous KH 2PO4.
4. The method for preparing a liquid quality control product of glycosylated hemoglobin according to claim 1, wherein: the volume ratio of the lower layer red blood cells to the buffer solution in the step 2) is 1:8-5:1.
5. The method for preparing a liquid quality control product of glycosylated hemoglobin according to claim 1, wherein: in the step 2), the rotation speed of the centrifugal machine is the same as that of the centrifugal machine in the step 1), and the centrifugal washing is carried out for 2-5 times in the step 2).
6. The method for preparing a liquid quality control product of glycosylated hemoglobin according to claim 1, wherein: in the step 3), the preservation solution comprises the following specific components: 5.0-10.0 g/L sodium citrate dihydrate, 0.4-0.8 g/L citric acid monohydrate, 16-22 g/L glucose, 5.0-8.0 g/L NaCl.
7. The method for preparing a liquid quality control product of glycosylated hemoglobin according to claim 1, wherein: in the step 3), the volume ratio of the red blood cells to the preservation solution is 10:1-1:10.
8. A liquid quality control product of glycosylated hemoglobin, characterized in that: the liquid quality control material of glycosylated hemoglobin according to any one of claims 1 to 7.
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN118150844A true CN118150844A (en) | 2024-06-07 |
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