CN109307769B - Preparation method of 12mmol/L blood glucose solution - Google Patents
Preparation method of 12mmol/L blood glucose solution Download PDFInfo
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Abstract
A preparation method of a 12mmol/L blood sugar solution comprises the following steps: 3L of phosphate buffer diluent is filled in a 5L measuring cup, the measuring cup is placed on a magnetic stirrer, and a stirring rotor rotates at a constant speed, wherein the adding amount mass concentration of each component in the phosphate buffer diluent is as follows: 0.055 percent of potassium dihydrogen phosphate, 0.237-0.238 percent of disodium hydrogen phosphate dihydrate, 0.15 percent of potassium chloride, 0.019-0.020 percent of sodium nitrate, 0.038-0.040 percent of disodium edetate dihydrate, 0.005-0.0055 percent of TritonX-100 and 0.05-0.06 percent of K40, and ultrapure water is used as a solvent. The configuration method of the invention does not generate a large amount of foam in the configuration process; and the preparation process of the 12mmol/L calibration solution is simple to operate, the efficiency is greatly improved, and the accuracy and precision of the calibration solution are easy to control.
Description
Technical Field
The invention relates to an in-vitro diagnostic reagent for medical instruments.
Background
The 12mmol/L blood sugar calibration solution is the most widely used calibration solution in biochemical research, the main components of the calibration solution are anhydrous glucose, Na2HPO4.2H2O, KH2PO4, NaNO3, KCL, K40 bactericides and the like, the balance is ultrapure water, the calibration solution is used as an in vitro diagnostic reagent 12mmol/L calibration solution, the researched technical indexes mainly relate to accuracy and precision, according to related technologies, the calibration solution with the accuracy of +/-10 percent and the precision of not more than 1.5 percent is found to be suitable for POCT blood sugar analyzers of various types in the market, and the test effect is superior to that of reagents matched with some blood sugar meters.
A12 mmol/L blood glucose calibration solution of a glucometer on the market generally adopts a solvent as injection water as the solvent, the pH value of the injection water is 5.0-7.0, and the conductivity is 1.3us/cm, the calibration solution produced by the method has more clinical adverse reactions and poor stability due to larger conductivity, and the technical indexes of the water quality used by the prepared calibration solution are that the pH value is 6.0-7.0, the water quality is closer to neutrality, and the conductivity is 0.65us/cm, so that the influence of microorganisms on the quality of the calibration solution is avoided. Meanwhile, the diluent used for preparing the calibration solution contains 5 times of Parmetol K40, so that the possibility of long-term stability of the calibration solution is ensured, and the result of blood test on an instrument is very optimistic.
The technical index accuracy of the calibration solution prepared by the method is +/-5%, the precision is less than or equal to 1.2%, the calibration solution is not easily influenced by the air environment during instrument detection, and the detection result is more accurate and reliable, so that the calibration solution required by testing of some full-automatic blood glucose/lactic acid analyzers in the market can be obtained through a large amount of experimental verification data on the addition amount of each chemical raw material in the concentrated solution.
Disclosure of Invention
The invention provides a preparation method of a 12mmol/L blood sugar solution, which comprises the following steps:
(1) 3L of phosphate buffer diluent is filled in a 5L measuring cup, the measuring cup is placed on a magnetic stirrer, and a stirring rotor rotates at a constant speed, wherein the addition mass concentration of each component in the phosphate buffer diluent is as follows: 0.055% of potassium dihydrogen phosphate, 0.237% -0.238% of disodium hydrogen phosphate dihydrate, 0.15% of potassium chloride, 0.019% -0.020% of sodium nitrate, 0.038% -0.040% of disodium edetate dihydrate, 0.005% -0.0055% of TritonX-100 and 0.05% -0.06% of K40, and using ultrapure water as a solvent; wherein TritonX-100 is triton-100, and the molecular formula is as follows: (C14H22O (C2H4O) n) in an amount of 0.005-0.0055% to achieve solubilization effect in the mixed solution, and K40 (parmetol K40 aqueous chloromethyl/methylisothiazolinone (CMI/MI) mixture in an amount of 0.05-0.06% to achieve sterilization effect;
(2) then slowly adding 108.09g of beta-D (+) anhydrous glucose into a 5L measuring cup, and continuously stirring on a magnetic stirrer for about 30 min;
(3) after the composition is completely dissolved, adding phosphate buffer diluent into a 5L measuring cup to the scale mark;
(4) Transferring 500ml of the mixed solution into a new 5L measuring cup, adding phosphate buffer diluent to the position of a 5L scale mark, continuously stirring on a magnetic stirrer for 30min, and standing for 30 min;
(5) transferring 100ml of phosphate buffer diluent into a new 5L measuring cup, continuously and slowly adding the phosphate buffer diluent to the 5L scale mark, continuously stirring for 30min, and standing for 30 min;
(6) after the solution is uniformly dissolved, obtaining a solution before filling, putting a 0.22um acetate fiber filter membrane soaked in triple distilled water into a filter, and filtering for three times to obtain the required calibration solution;
(7) if the solution is clear and transparent, sampling and detecting, performing three batches continuously, starting to test the accuracy of the solution, and if precipitation and turbidity phenomena exist, removing the precipitates and the turbidity phenomena and reconfiguring;
(8) and after the test is qualified, performing an on-machine verification test by using the 12mmol/L calibration solution, performing a calibration test on a blood sample by using a glucose/lactic acid analyzer and a blood glucose analyzer, and comparing and clinically verifying the consistency with a standard calibration product.
The invention has the following advantages:
1. a large amount of foam is not generated in the preparation process;
2. the preparation process of the 12mmol/L calibration solution is simple to operate, the efficiency is greatly improved, and the accuracy and the precision of the calibration solution are easy to control;
3. The repeatability is within +/-1%;
4. the 12mmol/L calibration solution has good stability and is not easily influenced by air;
5. can be used for the calibration of POCT glucometers of various brands, has good calibration effect, and simultaneously saves the use cost of the total reagent
Further, the present invention includes:
1. the test of the calibration solution on instruments such as a blood glucose meter and the like can be realized more accurately,
2. is convenient for simple and visual configuration in production,
3. the invention adopts a low-rotating-speed stirring and dissolving method, the prepared glucose is stirred for 30min, the anhydrous glucose belongs to a hydrophilic substance, the dissolution is easy, the uniformity of the solution is not influenced, the generation of foam and bubbles caused by high rotating speed is avoided, the low rotating speed is energy-saving, the method is suitable for the industrial production requirement,
4. the addition of K40 with 5 times of content is easy to control the stability of the calibration solution, so that the test result is more accurate
5. The ultrapure water is used as a solvent of the phosphate buffer diluent, so that the accuracy and precision of the preparation of the calibration solution are improved.
Detailed Description
The technical solution of the present invention is further explained by the following embodiments. It is to be understood that the specific embodiments described herein are merely illustrative of the invention and are not limiting of the invention.
The stage of the invention aims to:
Designing a technical formula containing 5 times of Parmetol K40 diluent in the first stage;
in the second stage, 12mmol/L of blood glucose calibration solution is prepared;
establishing a mature process flow according to the formula;
in the fourth stage, the blood glucose meter and other instruments are used for accuracy test, precision test, repeated test and clinical calibration consistency test, and the blood glucose meter and other instruments are put into production formally.
The final purpose is as follows: a method for preparing a 12mmol/L blood sugar calibration solution for whole blood sample detection.
1. The concentrates of this study have the following accessories:
(1) analytical balance (0.0001g)
(1) A magnetic stirrer;
(2) a digital temperature controller;
(3) a pH meter;
(4) a conductivity meter;
(5) a 5L large measuring cup;
(6) the prepared phosphate buffer diluent is placed in a small 100ml beaker;
(7) after the PH meter and the conductivity meter are calibrated, inserting electrodes of the PH meter and the conductivity meter into the solution;
(8) pressing the 'reading' buttons of the PH meter and the conductivity meter respectively;
(9) waiting for 1min-2min, and recording the reading on the PH meter and the reading on the conductivity;
(10) preparing 120 mmol/L10 times blood glucose calibration solution with the prepared buffer diluent, and then
Diluting for 10 times to obtain 12mmol/L calibration solution, diluting for 50 times to obtain sample,
making technical comparison with standard calibration solution and clinical test analysis
(1) The additive concentration and the additive amount of each chemical in 5L of phosphate buffer diluent are that the additive amount of potassium dihydrogen phosphate is 0.055%, the additive amount of disodium hydrogen phosphate dihydrate is 0.237% -0.238%, the additive amount of potassium chloride is 0.15%, the additive amount of sodium nitrate is 0.019% -0.020%, the additive amount of disodium edetate dihydrate is 0.038% -0.040%, the additive amount of TritonX-100 is 0.005% -0.0055%, the additive amount of K40 is 0.05% -0.06%, and ultrapure water is used as a solvent;
(2) dissolving all the inorganic salts mentioned above should be prepared beforehand for preliminary dissolution of 60% of the volume of the solvent, in order to avoid the generation of foam;
(3) for organic substances K40 and TritonX-100, one plays a role in sterilization, the other disperses cells in a blood sample to facilitate the detection of an instrument, and simultaneously reduces the surface tension of diluent to reduce the generation of bubbles and eliminate the interference of the bubbles on measurement, and the substance also has the function of sheath fluid; the adding sequence is that TritonX-100 is added first and then K40 is added;
(4) accurately weighing 108.09g of beta-D (+) anhydrous glucose, pouring the weighed beta-D (+) anhydrous glucose into a 5L measuring cup which is pre-filled with 3L of phosphate buffer diluent containing 5 times of K40 bactericide, continuously stirring for about 30min, and adding the phosphate buffer diluent to the position of 5L scale mark;
(5) Transferring 500ml of 120mmol/L concentrated calibration solution into a new 5L measuring cup, diluting by 10 times to obtain a 5L scale mark, continuously stirring on a magnetic stirrer for about 30min, and standing for 30 min;
(6) transferring 100ml of 12mmol/L calibration solution to a new 5L measuring cup, continuously and slowly adding phosphate buffer diluent to the position of a 5L scale mark, continuously stirring for about 30min, and standing for 30 min;
(7) the invention adopts a low-rotating-speed stirring and dissolving method, the prepared beta-D (+) anhydrous glucose belongs to hydrophilic substances and is easy to dissolve, the homogeneity of the solution is not influenced, and meanwhile, the low-rotating-speed energy-saving method is low in rotating speed and suitable for industrial production requirements;
(8) the content of K40 is 0.05% -0.06%, the prepared calibration solution has better stability, and the tested result is more accurate;
(9) the water quality is preferably triple distilled water or ultrapure water
3. The preparation method comprises the following steps:
(1) 3L of phosphate buffer diluent is filled in a 5L measuring cup (the volume needs to be calibrated), the measuring cup is placed on a magnetic stirrer, and a stirring rotor rotates at a constant speed;
(2) then 108.09g of beta-D (+) anhydrous glucose is slowly added into a 5L measuring cup, and the mixture is placed on a magnetic stirrer to be continuously stirred for about 30 min;
(3) after the chemicals are completely dissolved, adding phosphate buffer diluent into a 5L measuring cup to the scale mark;
(4) Transferring 500ml of the mixed solution into a new 5L measuring cup, adding phosphate buffer diluent to the position of a 5L scale mark, continuously stirring on a magnetic stirrer for about 30min, and standing for 30 min;
(5) transferring 100ml of phosphate buffer diluent into a new 5L measuring cup, continuously and slowly adding the phosphate buffer diluent to the position of the L5L scale mark, continuously stirring for 30min, and standing for 30 min;
(6) dissolving uniformly to obtain a solution before filling, putting a 0.22um acetate fiber filter membrane soaked in triple distilled water into a filter, and filtering for three times to obtain the required calibration solution;
(7) if the solution is clear and transparent, sampling and detecting, continuously making three batches, starting to test the accuracy of the solution, and if precipitation and turbidity phenomena exist, removing the solution and reconfiguring;
(8) and after the test is qualified, performing an on-machine verification test by using the 12mmol/L calibration solution, performing a calibration test on a blood sample by using a glucose/lactic acid analyzer and a blood glucose analyzer, and comparing and clinically verifying the consistency with a standard calibration product.
Example one
1. Putting 3L of phosphate buffer diluent into a 5L measuring cup (the volume needs to be calibrated and is calibrated), putting the measuring cup under a stirrer, extending a stirring rod to 1/2 parts of the liquid level, stirring at a constant speed, continuously adding anhydrous glucose into the 5L measuring cup, completely dissolving chemicals, then adding phosphate buffer diluent into the 5L measuring cup to a scale mark (a calibrated scale mark), continuously stirring, stirring for 30min, standing for 30min, transferring 500ml of mixed solution into a new 5L measuring cup, continuously adding phosphate buffer diluent to the scale mark, stirring for 30min, standing for 30min, transferring 100ml of phosphate buffer diluent into the new 5L measuring cup, continuously adding phosphate buffer diluent to the 5L scale mark, continuously stirring for 30min, standing for 30min, sterilizing and filtering the solution after finishing, thus obtaining the required 12mmol/L blood glucose calibration solution,
2. Standing for 1h to obtain solution before filling, testing pH value and conductivity of the obtained concentrated solution if the obtained concentrated solution is clear and transparent and has no precipitation and turbidity, diluting into system solution after the concentrated solution is qualified, performing on-machine test, sampling and detecting, and performing three batches continuously, wherein the batches are 20161208, 20161212 and 20161214 respectively;
3. and (4) conclusion:
example two
1. Putting 3L of phosphate buffer diluent into a 5L measuring cup (the volume needs to be calibrated and is calibrated), putting the measuring cup under a stirrer, extending a stirring rod to 1/2 parts of the liquid level, stirring at a constant speed, continuously adding anhydrous glucose into the 5L measuring cup, completely dissolving chemicals, then adding phosphate buffer diluent into the 5L measuring cup to a scale mark (a calibrated scale mark), continuously stirring, stirring for 30min, standing for 30min, transferring 500ml of mixed solution into a new 5L measuring cup, continuously adding phosphate buffer diluent to the scale mark, stirring for 30min, standing for 30min, transferring 100ml of phosphate buffer diluent into the new 5L measuring cup, continuously adding phosphate buffer diluent to the 5L scale mark, continuously stirring for 30min, standing for 30min, sterilizing and filtering the solution after finishing, thus obtaining the required 12mmol/L blood glucose calibration solution,
2. Standing for 1h to obtain solution before filling, testing pH value and conductivity of the obtained concentrated solution if the obtained concentrated solution is clear and transparent and has no precipitation and turbidity, diluting into system solution after the concentrated solution is qualified, performing on-machine test, sampling and detecting, and performing three batches continuously, wherein the batches are 20161208, 20161212 and 20161214 respectively;
3. and (4) conclusion:
EXAMPLE III
1. Filling 3L of phosphate buffer diluent into a 5L measuring cup (the volume needs to be calibrated and is calibrated), placing the measuring cup under a stirrer, extending a stirring rod into the liquid level 1/2, stirring at a constant speed, then, continuously adding anhydrous glucose into the 5L measuring cup, after chemical products such as the anhydrous glucose and the like are completely dissolved, then adding phosphate buffer diluent into the 5L measuring cup to the position of a scale mark (the calibrated scale mark), continuously stirring, stirring for 30min, standing for 30min, transferring 500ml of mixed solution into a new 5L measuring cup, continuously adding phosphate buffer diluent to the scale mark, stirring for 30min, standing for 30min, transferring 100ml of phosphate buffer diluent to a new 5L measuring cup, continuously adding phosphate buffer diluent to the position of a 5L scale mark, continuously stirring for 30min, standing for 30min, and sterilizing and filtering the solution after the stirring is finished to obtain the required blood sugar calibration solution of 12 mmol/L;
2. Standing for 1h to obtain solution before filling, testing pH value and conductivity of the obtained concentrated solution if the obtained concentrated solution is clear and transparent and has no precipitation and turbidity, diluting into system solution after the concentrated solution is qualified, performing on-machine test, sampling and detecting, and performing three batches continuously, wherein the batches are 20161208, 20161212 and 20161214 respectively;
3. and (4) conclusion:
citation: YYT 0456.1-2014 reagent part 1 for hematology analyzers: cleaning fluid; YYT 0456.3-2014 reagent part 3 for hematology analyzers: a diluent; ISO18113-2 part 2: professional in vitro diagnostic reagents; part 1 of the standard GBT1.1-2009 standardization work guide rule for GBT 26124-2011 clinical chemistry in-vitro diagnostic kit standard (GB) YYT 0701-2008 blood cell analyzer: standard construction and writing (Abstract)
The present invention is not limited to the above embodiments, and any modifications, equivalent substitutions, improvements, etc. within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Claims (1)
1. A preparation method of a 12mmol/L blood sugar solution is characterized by comprising the following steps:
(1) 3L of phosphate buffer diluent is filled in a 5L measuring cup, the measuring cup is placed on a magnetic stirrer, and a stirring rotor rotates at a constant speed, wherein the adding amount mass concentration of each component in the phosphate buffer diluent is as follows: 0.055% of potassium dihydrogen phosphate, 0.237% -0.238% of disodium hydrogen phosphate dihydrate, 0.15% of potassium chloride, 0.019% -0.020% of sodium nitrate, 0.038% -0.040% of disodium edetate dihydrate, 0.005% -0.0055% of TritonX-100 and 0.05% -0.06% of K40, and using ultrapure water as a solvent;
(2) Then 108.09g of beta-D (+) anhydrous glucose is slowly added into a 5L measuring cup, and the mixture is placed on a magnetic stirrer to be continuously stirred for about 30 min;
(3) after the composition is completely dissolved, adding phosphate buffer diluent into a 5L measuring cup to the scale mark;
(4) transferring 500ml of the mixed solution into a new 5L measuring cup, adding phosphate buffer diluent to the position of a 5L scale mark, continuously stirring on a magnetic stirrer for 30min, and standing for 30 min;
(5) transferring 100ml of phosphate buffer diluent into a new 5L measuring cup, continuously and slowly adding the phosphate buffer diluent to the 5L scale mark, continuously stirring for 30min, and standing for 30 min;
(6) after the solution is uniformly dissolved, obtaining a solution before filling, putting a 0.22um acetate fiber filter membrane soaked in triple distilled water into a filter, and filtering for three times to obtain the required calibration solution;
(7) if the solution is clear and transparent, sampling and detecting, continuously making three batches, starting to test the accuracy of the solution, and if precipitation and turbidity phenomena exist, removing the solution and reconfiguring;
(8) and after the test is qualified, performing an on-machine verification test by using the 12mmol/L calibration solution, performing a calibration test on a blood sample by using a glucose/lactic acid analyzer and a blood glucose analyzer, and comparing and clinically verifying the consistency with a standard calibration product.
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