阅读说明：本技术 一种抗凝血酶iii测定试剂盒 (Antithrombin III determination kit ) 是由 焦东燕 刘芳 王玉玉 于 2020-08-28 设计创作，主要内容包括：本发明涉及体外诊断技术领域,尤其涉及一种抗凝血酶III测定试剂盒,解决了现有技术中存在的测定试剂盒的测定速率较慢,且在测定完成后试剂反应仍在继续进行中,影响测定实验结果数值,导致最终结果出现偏差的缺点,包括试剂一、试剂二、提速液及终止液；试剂一包括凝血酶、肝素、缓冲液及盐；试剂二包括凝血酶的发色底物、缓冲液及甘露醇；提速液包括磷酸氢二钠溶液；终止液包括聚氯乙烯树酯及叠氮化钠。本发明通过试剂一、试剂二、提速液及终止液组合成试剂盒,利用试剂一、试剂二、提速液快速实现抗凝血酶III的测定,在测定完成后利用终止液来停止反应,避免缓慢继续反应造成的数据偏差,整体测定试剂盒的使用方便,测定快捷,数据准确。(The invention relates to the technical field of in-vitro diagnosis, in particular to an antithrombin III determination kit, which solves the defects that the determination rate of the determination kit in the prior art is slow, and the reagent reaction is still in the process after the determination is finished, so that the numerical value of the determination experiment result is influenced, and the final result is deviated; reagent one comprises thrombin, heparin, buffer solution and salt; the reagent II comprises a thrombin chromogenic substrate, a buffer solution and mannitol; the accelerating solution comprises a disodium hydrogen phosphate solution; the stop solution comprises polyvinyl chloride resin and sodium azide. According to the kit, the reagent I, the reagent II, the accelerating solution and the stopping solution are combined to form the kit, the determination of the antithrombin III is rapidly realized by using the reagent I, the reagent II and the accelerating solution, the reaction is stopped by using the stopping solution after the determination is finished, the data deviation caused by slow continuous reaction is avoided, and the whole determination kit is convenient to use, rapid in determination and accurate in data.)

1. An antithrombin III determination kit is characterized by comprising a reagent I, a reagent II, a speed-up solution and a stop solution;

the reagent I comprises thrombin, heparin, a buffer solution and salt;

the reagent II comprises a thrombin chromogenic substrate, a buffer solution and mannitol;

the accelerating solution comprises a disodium hydrogen phosphate solution;

the stop solution comprises polyvinyl chloride resin and sodium azide.

2. The antithrombin III assay kit according to claim 1, wherein in reagent one: the concentration of thrombin is 5-20 IU/ml, the content of heparin is 0.8-1.2 wt%, and the content of salt is 0.25-0.5 wt%.

3. The antithrombin III assay kit according to claim 2, wherein in reagent two: the content of the thrombin chromogenic substrate is 2.5-4.5 wt%, and the content of mannitol is 0.3-0.5 wt%.

4. The antithrombin III assay kit as claimed in claim 3, wherein the concentration of the disodium hydrogen phosphate solution in the accelerating solution is 1 to 3IU/ml, and in the stop solution: the content of the polyvinyl chloride resin is 0.3-1.0 wt% and the content of the sodium azide is 0.5-0.8 wt%.

5. The antithrombin III assay kit according to any one of claims 1 or 4, wherein the chromogenic substrate is one of S2238, S2160 and Chromozym TH.

6. The antithrombin III assay kit according to any one of claims 1 or 4, wherein the buffer is Tris.

7. The antithrombin III assay kit according to any one of claims 1 or 4, wherein the salt is one or more of sodium chloride, potassium chloride, magnesium chloride, ammonium sulfate.

8. A method of using the antithrombin III assay kit according to any one of claims 1 to 7, comprising the steps of:

s1: placing the liquid to be detected in a first reagent bottle, a second reagent bottle and a third reagent bottle;

s2: adding a first reagent into the first reagent bottle, the second reagent bottle and the third reagent bottle respectively, and incubating for 5-10 min;

s3: adding a second reagent and a speed-increasing solution into the first reagent bottle, the second reagent bottle and the third reagent bottle respectively, mixing uniformly, and incubating for 5-10 min;

s4: and adding the stop solution into the first reagent bottle, the second reagent bottle and the third reagent bottle respectively, observing the absorbance of the first reagent bottle, the second reagent bottle and the third reagent bottle, and comparing the absorbance with standard solution with known concentration to obtain the antithrombin III concentration in the detection solution.

9. The method of using the antithrombin III assay kit according to claim 8, wherein the amount of the first reagent is 2.5 to 5.8% of the volume of the detection solution, the amount of the second reagent is 1.3 to 2.5% of the volume of the detection solution, the amount of the accelerating solution is 0.5 to 1.2% of the volume of the detection solution, and the amount of the stop solution is 0.5 to 1.4% of the volume of the detection solution.

Technical Field

The invention relates to the technical field of in-vitro diagnosis, in particular to an antithrombin III determination kit.

Background

According to the principle of naming medical instruments, the names of products are unified, and the names of products on the market at present are divided into two types, namely an antithrombin III determination kit (chromogenic substrate method) and an antithrombin determination kit (chromogenic substrate method).

The international standard for antithrombin was prepared and assigned by the national institute for biological products (NIBSC) of the uk. Based on the information provided by this agency, the first generation of international reference (72/1) established in 1978 by the international health organization (WHO) under the designation antithrombin iii. The nomenclature follows the publication prothrombin activated antithrombin reaction am.j. physiol.176(1) of seegers wh, johnson jf, FellC (1954): 97-103. In this article there are four references to antithrombin, the name antithrombin III which we now refer to as antithrombin. The International Society for Thrombosis and Hemostasis (ISTH) has now termed antithrombin, and no longer uses antithrombin iii. The second generation international reference (93/578) prepared by NIBSC and established by WHO in 1994, and the third generation (08/258) international reference established in 2010, have therefore no longer used the name "antithrombin iii" but rather the name "antithrombin".

Although the nomenclature, international reference, has been unified internationally, the nomenclature of the antithrombin III assay kit (chromogenic substrate method) is used herein according to domestic registration. Antagonistic to the function of the coagulation system in the human body is the anticoagulant system, and under normal conditions, the two systems are in dynamic equilibrium, thereby maintaining normal blood flow in the blood vessel. Antithrombin III is the most important component in an anticoagulation system, is synthesized by liver, is a multifunctional serine protease inhibitor, can inhibit thrombin generation, has strong anticoagulation activity and accounts for 70% of the activity of plasma anticoagulation enzyme, is mostly measured by a chromogenic substrate method, has slow measurement rate in the conventional measurement kit, and has influence on the numerical value of a measurement experiment result after the reagent reaction is still carried out after the measurement is finished, so that the deviation of the final result is caused.

Therefore, we propose an antithrombin III assay kit to solve the above problems.

Disclosure of Invention

The invention aims to solve the defects that the determination speed of a determination kit in the prior art is slow, and the reagent reaction is still in the process after the determination is finished, so that the numerical value of the determination experiment result is influenced, and the final result is deviated.

In order to achieve the purpose, the invention adopts the following technical scheme:

the invention provides an antithrombin III determination kit, which comprises a reagent I, a reagent II, a speed-up solution and a stop solution;

the reagent I comprises thrombin, heparin, a buffer solution and salt;

the reagent II comprises a thrombin chromogenic substrate, a buffer solution and mannitol;

the accelerating solution comprises a disodium hydrogen phosphate solution;

the stop solution comprises polyvinyl chloride resin and sodium azide.

Preferably, in the reagent one: the concentration of thrombin is 5-20 IU/ml, the content of heparin is 0.8-1.2 wt%, and the content of salt is 0.25-0.5 wt%.

Preferably, in the reagent two: the content of the thrombin chromogenic substrate is 2.5-4.5 wt%, and the content of mannitol is 0.3-0.5 wt%.

Preferably, the concentration of the disodium hydrogen phosphate solution in the accelerating solution is 1-3 IU/ml, and in the stopping solution: the content of the polyvinyl chloride resin is 0.3-1.0 wt% and the content of the sodium azide is 0.5-0.8 wt%.

Preferably, the chromogenic substrate is one of S2238, S-2160 and Chromozym TH.

Preferably, the buffer is Tris.

Preferably, the salt is one or more of sodium chloride, potassium chloride, magnesium chloride and ammonium sulfate.

Preferably, the use method of the antithrombin III determination kit comprises the following steps:

s1: placing the liquid to be detected in a first reagent bottle, a second reagent bottle and a third reagent bottle;

s2: adding a first reagent into the first reagent bottle, the second reagent bottle and the third reagent bottle respectively, and incubating for 5-10 min;

s3: adding a second reagent and a speed-increasing solution into the first reagent bottle, the second reagent bottle and the third reagent bottle respectively, mixing uniformly, and incubating for 5-10 min;

s4: and adding the stop solution into the first reagent bottle, the second reagent bottle and the third reagent bottle respectively, observing the absorbance of the first reagent bottle, the second reagent bottle and the third reagent bottle, and comparing the absorbance with standard solution with known concentration to obtain the antithrombin III concentration in the detection solution.

Preferably, the adding amount of the first reagent is 2.5-5.8% of the volume of the detection liquid, the adding amount of the second reagent is 1.3-2.5% of the volume of the detection liquid, the adding amount of the accelerating liquid is 0.5-1.2% of the volume of the detection liquid, and the adding amount of the stop solution is 0.5-1.4% of the volume of the detection liquid.

Compared with the prior art, the invention has the beneficial effects that:

1. according to the invention, the speed-up solution is arranged in the determination kit, the non-traditional disodium hydrogen phosphate solution is utilized to improve the determination reaction rate and accelerate the obtaining of experimental results, and meanwhile, the disodium hydrogen phosphate solution can also play a role of a quality modifier, so that the reagent has multiple effects, and the reaction determination can be stably carried out while the determination reaction rate is improved.

2. The invention sets the stop solution in the determination kit, and utilizes the polyvinyl chloride resin and the sodium azide to stop the reaction of the reagent in time, thereby avoiding the influence on the actual determination data value caused by the continuous reaction after the determination is finished, wherein the polyvinyl chloride resin has wide sources, convenient and stable preparation and low cost, and the cost of the determination kit is reduced.

3. According to the invention, the reagent I, the reagent II, the accelerating solution and the stopping solution are combined to form the measuring kit, the antithrombin III is measured normally at a high speed by using the reagent I, the reagent II and the accelerating solution, and the reaction is stopped in time by using the stopping solution after the measurement is finished, so that the data deviation caused by slow continuous reaction is avoided, and the whole measuring kit is convenient to use, rapid to measure and accurate in data.

Detailed Description

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. In case of conflict, the present specification, including definitions, will control. When "mass, concentration, temperature, time, or other value or parameter is expressed as a range, preferred range, or as a range defined by a list of upper preferable values and lower preferable values, this is to be understood as specifically disclosing all ranges formed from any pair of any upper range limit or preferred value and any lower range limit or preferred value, regardless of whether ranges are separately disclosed. For example, a range of 1 to 50 should be understood to include any number, combination of numbers, or subrange selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50, and all fractional values between the above integers, e.g., 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, and 1.9. With respect to sub-ranges, specifically consider "nested sub-ranges" that extend from any endpoint within the range. For example, nested sub-ranges of exemplary ranges 1-50 may include 1-10, 1-20, 1-30, and 1-40 in one direction, or 50-40, 50-30, 50-20, and 50-10 in another direction. "

The present invention is further illustrated below with reference to specific examples in which various processes and methods not described in detail are conventional methods well known in the art. Materials, reagents, devices, instruments, apparatuses and the like used in the following examples are commercially available unless otherwise specified.