阅读说明：本技术 谷胱甘肽还原酶测定试剂校准品及制备方法 (Glutathione reductase determination reagent calibrator and preparation method thereof ) 是由 万蒙 周惠良 于 2019-09-29 设计创作，主要内容包括：本发明公开了一种谷胱甘肽还原酶测定试剂校准品,包括以下组分的冻干品：磷酸盐缓冲液50～80份,草酸钾2～5份,牛血清5～10份,纯品谷胱甘肽还原酶0.1～0.5份,蔗糖2～5份,防腐剂1～2份。磷酸盐缓冲液能便于校准品的制备成型,防腐剂配合草酸钾、牛血清能延长校准品的质量,蔗糖能防止纯品谷胱甘肽还原酶在制备过程中失活。本发明还公开了一种谷胱甘肽还原酶测定试剂校准品的制备方法,包括以下步骤：取纯品谷胱甘肽还原酶,加入所述磷酸盐缓冲液稀释,得稀释液；向稀释液中加入草酸钾溶液和所述牛血清,保温；加入蔗糖溶液,混合均匀,静置,得冻干材料；将冻干材料进行预冷冻,得初始冻干品；将初始冻干品进行真空冷冻干燥,得谷胱甘肽还原酶测定试剂校准品。(The invention discloses a glutathione reductase assay reagent calibrator, which comprises a freeze-dried product of the following components: 50-80 parts of phosphate buffer solution, 2-5 parts of potassium oxalate, 5-10 parts of bovine serum, 0.1-0.5 part of pure glutathione reductase, 2-5 parts of cane sugar and 1-2 parts of preservative. The phosphate buffer solution can facilitate the preparation and the forming of the calibrator, the preservative can prolong the quality of the calibrator by matching with potassium oxalate and bovine serum, and the sucrose can prevent the inactivation of the pure glutathione reductase in the preparation process. The invention also discloses a preparation method of the glutathione reductase determination reagent calibrator, which comprises the following steps: taking a pure glutathione reductase, and adding the pure glutathione reductase into the phosphate buffer solution for dilution to obtain a diluent; adding a potassium oxalate solution and the bovine serum into the diluent, and preserving heat; adding sucrose solution, mixing, and standing to obtain lyophilized material; pre-freezing the freeze-dried material to obtain an initial freeze-dried product; and (4) carrying out vacuum freeze drying on the initial freeze-dried product to obtain the glutathione reductase assay reagent calibrator.)

1. A glutathione reductase assay reagent calibrator is characterized by comprising a freeze-dried product of the following components:

50-80 parts of phosphate buffer solution,

2-5 parts of potassium oxalate,

5-10 parts of bovine serum,

0.1-0.5 part of pure glutathione reductase,

2-5 parts of cane sugar,

1-2 parts of a preservative.

2. The glutathione reductase assay reagent calibrator according to claim 1, wherein the phosphate buffer comprises disodium hydrogen phosphate and sodium dihydrogen phosphate in a mass ratio of 2:3 to 6.

3. The glutathione reductase assay reagent calibrator according to claim 1, wherein the preservative comprises sodium azide and sodium glutamate in a mass ratio of 3: 1-2.

4. The method for producing a glutathione reductase assay reagent calibrator according to any one of claims 1 to 3, comprising the steps of:

taking the pure glutathione reductase, and adding a phosphate buffer solution for dilution to obtain a diluent;

adding a potassium oxalate solution and the bovine serum into the diluent, and preserving heat for 2 hours;

adding sucrose solution, mixing, and standing to obtain lyophilized material;

pre-freezing the freeze-dried material to obtain an initial freeze-dried product;

and carrying out vacuum freeze drying on the initial freeze-dried product to obtain the glutathione reductase determination reagent calibrator.

5. The method according to claim 4, wherein the concentration of the phosphate buffer is 50 to 400mmol/L, the concentration of the potassium oxalate solution is 100 to 200mmol/L, and the concentration of the bovine serum is 30 to 95%.

6. The method according to claim 4, wherein the concentration of glutathione reductase in the diluted solution is 10 to 100U/L.

7. The preparation method according to claim 4, wherein the step of adding a potassium oxalate solution to the diluent and maintaining the temperature for 12 hours is carried out at a temperature of 15 to 25 ℃.

8. The method according to claim 4, wherein the step of adding the sucrose solution, mixing uniformly, and standing comprises the steps of:

and placing the mixture in an environment with the temperature of 2-8 ℃ for 24 hours.

9. The method according to claim 4, wherein the step of pre-freezing the lyophilized material to obtain an initial lyophilized product comprises:

keeping the freeze-dried material in a freezing environment at-30 to-40 ℃ for 4 to 8 hours;

and gradually cooling the freezing environment, wherein the cooling range is 2-3 ℃/s until the temperature of the freezing environment is-60 to-70 ℃, and keeping for 10-15 hours.

10. The method of claim 4, wherein the step of vacuum freeze-drying the initial lyophilizate comprises:

placing the initial freeze-dried product in an environment with the atmospheric pressure of 5Pa, wherein the environment temperature is-40 to-60 ℃, and keeping for 24 to 36 hours;

and placing the initial freeze-dried product in an environment with the atmospheric pressure of 15Pa, wherein the environment temperature is-20 to-30 ℃, and keeping for 8 to 12 hours.

Technical Field

The invention relates to the technical field of biochemical detection, in particular to a glutathione reductase determination reagent calibrator and a preparation method thereof.

Background

Glutathione Reductase (GR) is contained in blood, and its content is measured by a glutathione reductase measuring reagent.

In the process of producing the glutathione reductase assay reagent, a calibrator is needed to be used for calibrating the glutathione reductase assay reagent, the value of the calibrator is used for a reference substance of an independent variable in a calibration function, and the calibrator has a definite value and known measurement uncertainty, and the aim is to calibrate a certain measurement system so as to establish the metrological traceability of the measurement result of the system. At present, the calibrator mostly adopts liquid pure glutathione reductase, buffer solution and stabilizer, and the calibrator is used as standard glutathione reductase to detect whether the glutathione reductase determination reagent is qualified or not and calibrate a calculation formula of the glutathione reductase determination reagent.

The existing glutathione reductase measuring reagent calibrator is in a liquid state, so that the product is difficult to store after being unsealed, and is discarded if not used, thereby causing resource waste.

Disclosure of Invention

An object of the present invention is to provide a calibration reagent for measuring glutathione reductase which has a long shelf life after unsealing.

A glutathione reductase assay reagent calibrator comprises a freeze-dried product of the following components:

50-80 parts of phosphate buffer solution,

2-5 parts of potassium oxalate,

5-10 parts of bovine serum,

0.1-0.5 part of pure glutathione reductase,

2-5 parts of cane sugar,

1-2 parts of a preservative.

The invention has the beneficial effects that: the phosphate buffer solution can facilitate the preparation and the forming of the calibrator, the preservative can prolong the quality of the calibrator by matching with potassium oxalate and bovine serum, and the sucrose can prevent the inactivation of the pure glutathione reductase in the preparation process.

In addition, the glutathione reductase assay reagent calibrator provided by the present invention may have the following additional technical features:

further, the phosphate buffer solution comprises disodium hydrogen phosphate and sodium dihydrogen phosphate, and the mass ratio of the disodium hydrogen phosphate to the sodium dihydrogen phosphate is 2: 3-6.

Further, the preservative comprises sodium azide and sodium glutamate in a mass ratio of 3: 1-2.

Another object of the present invention is to provide a method for preparing the glutathione reductase assay reagent calibrator, comprising the steps of:

taking the pure glutathione reductase, and adding the phosphate buffer solution for dilution to obtain a diluent;

adding a potassium oxalate solution and the bovine serum into the diluent, and preserving heat for 2 hours;

adding sucrose solution, mixing, and standing to obtain lyophilized material;

pre-freezing the freeze-dried material to obtain an initial freeze-dried product;

and carrying out vacuum freeze drying on the initial freeze-dried product to obtain the glutathione reductase determination reagent calibrator.

Further, the concentration of the phosphate buffer solution is 50-400 mmol/L, the concentration of the potassium oxalate solution is 100-200 mmol/L, and the concentration of the bovine serum is 30-95%.

Further, the concentration of the glutathione reductase in the diluent is 10-100U/L.

Further, adding a potassium oxalate solution into the diluent, and keeping the temperature for 12 hours at 15-25 ℃.

Further, in the step of adding a sucrose solution, uniformly mixing and standing, the step of standing is as follows:

and placing the mixture in an environment with the temperature of 2-8 ℃ for 24 hours.

Further, in the step of pre-freezing the freeze-dried material to obtain an initial freeze-dried product, the pre-freezing step is as follows:

keeping the freeze-dried material in a freezing environment at-30 to-40 ℃ for 4 to 8 hours;

and gradually cooling the freezing environment, wherein the cooling range is 2-3 ℃/s until the temperature of the freezing environment is-60 to-70 ℃, and keeping for 10-15 hours.

Further, the step of vacuum freeze-drying the initial lyophilizate comprises:

placing the initial freeze-dried product in an environment with the atmospheric pressure of 5Pa, wherein the environment temperature is-40 to-60 ℃, and keeping for 24 to 36 hours;

and placing the initial freeze-dried product in an environment with the atmospheric pressure of 15Pa, wherein the environment temperature is-20 to-30 ℃, and keeping for 8 to 12 hours.

Additional aspects and advantages of the invention will be set forth in part in the description which follows and, in part, will be obvious from the description, or may be learned by practice of the invention.

Detailed Description

In order to make the objects, features and advantages of the present invention comprehensible, specific embodiments accompanied with examples are described in detail below. Several embodiments of the invention are given in the examples. This invention may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete.