CN117778111A - Cleaning solution for thrombin-antithrombin complex (TAT) detection and preparation method thereof - Google Patents

Abstract

The application discloses a cleaning solution for thrombin-antithrombin complex (TAT) detection and a preparation method thereof, wherein the cleaning solution comprises the following components: inorganic salt, metal ion, buffering agent, pertinence complex stabilizer, monomer stabilizer, surfactant active agent and defoaming agent, pertinence complex stabilizer can improve thrombin-antithrombin complex (TAT) stability in plasma, be difficult for taking place the dissociation of complex along with the progress of wasing, combine other salt ion, surfactant active agent and saccharide composition also can reduce the monomer and form the possibility of new complex, through a large amount of realization proving surfactant active agent and pertinence complex stabilizer jointly use, TAT complex stability is better, the defoaming agent can prevent to produce the bubble in the washing liquid or washing process, thereby prevent the emergence of jump value phenomenon, improve detection accuracy and precision, consequently this washing liquid can reduce nonspecific interfering substance, reduce degradation or the denaturation of waiting to detect the target object.

Description

Cleaning solution for thrombin-antithrombin complex (TAT) detection and preparation method thereof

Technical Field

The application relates to the technical field of biological analysis, in particular to a cleaning solution for thrombin-antithrombin complex (TAT) detection and a preparation method thereof.

Background

After thrombin formation in vivo, thrombin has a half-life in blood of only a few seconds, direct measurement is difficult, thrombin moiety rapidly binds to Antithrombin (AT) to form thrombin-antithrombin-antithrombin complex, TAT, which is a molecular marker reflecting thrombin generation, sensitively reflects the degree of activation of the coagulation system, and directly reflects the initiation of the coagulation system. Elevated TAT can predict early thrombosis and risk of recurrence, and early DIC risk.

The current TAT detection method mainly comprises the steps of magnetic particle chemiluminescence detection, wherein the detection flow comprises loading, incubation, cleaning, chromogenic excitation and interpretation, the cleaning step needs 2-3 times, and the cost of the cleaning liquid can dissociate trace TAT compound product compound, so that the cleaning step plays a quite important role in the accuracy of a detection result, and the cleaning effect of the cleaning liquid is to remove non-target detection objects, stop the magnetic particles from continuously capturing detection target objects and reduce the influence of the non-target objects on the detection result. The cleaning solution not only can provide a proper buffer system environment for the antigen-antibody result in the detection reaction, but also can reduce the influence of other interfering substances on the subsequent detection reaction, and end the continuous combination of the antigen-antibody, and the surface active components in the cleaning solution can effectively separate the combined immunoreactants and nonspecific interfering substances, so that the magnetic particle chemiluminescence detection reaction can play a role in normal detection.

Furthermore, the pipeline and the sample injection needle in the magnetic particle chemiluminescence detection equipment are recycled, so that cross contamination can be generated in detection, and the stability of detection data is further affected. Therefore, the proper chemiluminescent cleaning liquid has important significance for maintaining a pipeline and a sample injection needle of the magnetic particle chemiluminescent detection equipment and ensuring the accuracy of sample testing.

The requirement for the cleaning solution may be higher for some special detection targets, especially for some active substances sensitive to the buffer system, if the buffer components of the cleaning solution are not suitable, degradation or denaturation of the target to be detected may be directly caused, and finally the accuracy and stability of the detection result of the detection reagent may be reduced.

The invention discloses a cleaning solution for chemiluminescent immunoassay, which is disclosed in the patent application number 201810130445X and is prepared from a surfactant, a non-protein protecting agent and a buffer agent, wherein the surfactant comprises an anionic surfactant, a polyoxyethylene ether type nonionic surfactant and an amide type nonionic surfactant, the molecular formula of the anionic surfactant is R1-SO3-M, wherein R1 is alkyl with 10-15 carbon atoms, and M is one of alkali metals. The repeatability and the specificity of the test reagent can be ensured, and the sensitivity of the test reagent can be improved.

The invention patent with the application number of 2017104520526 and the patent name of cleaning liquid discloses a cleaning liquid for chemiluminescence immunoassay, which comprises a phosphoric acid buffer system, inorganic salt, a surfactant A, a surfactant B and a preservative, wherein the phosphoric acid buffer system is one of a K2HPO4-KH2PO4 system or a Na2HPO4.12H2O-KH2PO4 system, the inorganic salt is one or two of NaCl or KCl, the surfactant A is one of Tween-20 or TritonX-100, the surfactant B is sodium cholate, the preservative is one or two of Proclin300 or Proclin950, the concentration of the phosphoric acid buffer system in the cleaning liquid is 0.1-0.5 mol/L based on phosphate, the inorganic salt content in the cleaning liquid is 90g/L-225g/L, the volume percentage of the surfactant A in the cleaning liquid is 1-2.5%, the content of the surfactant B in the cleaning liquid is 5g/L-25g/L, and the pH value in the cleaning liquid is 0.5-7.5% by volume percentage. The cleaning liquid has the advantages of universal system, good stability, long effective period, high accuracy, high precision and low cost, and is far longer than the commercial cleaning liquid.

However, none of the above-mentioned comparison documents discloses a technical scheme how the cleaning solution reduces the continuous combination of antigen and antibody, reduces non-specific interfering substances, and reduces degradation or denaturation of the target to be detected, thereby improving the accuracy and stability of the detection result.

Content of the application

Technical problem to be solved

Aiming at the defects of the prior art, the application provides a cleaning solution for thrombin-antithrombin complex (TAT) detection and a preparation method thereof, which can reduce nonspecific interfering substances and reduce degradation or denaturation of a target object to be detected, thereby improving the accuracy and stability of a detection result.

Technical proposal

In order to solve the technical problems, the specific technical scheme adopted by the application is as follows:

a wash solution for thrombin-antithrombin complex (TAT) detection, comprising: inorganic salts, metal ions, buffers, targeted complex stabilizers, monomer stabilizers, surfactants, and defoamers.

The stability of thrombin-antithrombin complex (TAT) in plasma can be improved by the targeted complex stabilizer in the cleaning liquid, the dissociation of the complex is not easy to occur along with the cleaning, the possibility of forming a new complex by combining other salt ions, surfactants and saccharide components can be reduced, the combination of the surfactants and the targeted complex stabilizer is proved by a large number of realizations, so that the TAT complex has better stability, bubbles generated in the process of adding the cleaning liquid or cleaning can be reduced by the defoamer, the occurrence of the jump value phenomenon is reduced, and the detection accuracy and precision are improved.

Preferably, the metal ion is provided by the inorganic salt, the inorganic salt can be sodium chloride or potassium chloride, the sodium ion or potassium ion is provided by sodium chloride or potassium chloride, the concentration of the metal ion is 100-500mM, the inorganic salt and the metal ion play a role in maintaining the surface structure of protein, and the cleaning solution can have certain ionic strength, so that the antibody can perform a binding function characteristically, and meanwhile, thrombin-antithrombin complex (TAT) can also exist stably.

Preferably, the pH value of the buffer is 7.0-7.5, the concentration of the buffer is 10-100mM, the buffer adopts Tris buffer, the buffer can ensure that the protein can be stably stored for a long time, the pH value of the buffer is regulated to 7.0-7.5 by adopting a solid acid regulator before use, and the activity of TAT and diagnostic antibodies can be well stabilized within the range of the buffer system, so that the detection result is more stable and reliable.

Preferably, the targeted compound stabilizer is at least one of mannitol, trehalose and arginine, the concentration of the targeted compound stabilizer is 0.1-0.5% w/v, the targeted compound stabilizer plays a role in stabilizing the compound, and the phenomenon of compound dissociation caused by cleaning can be reduced.

Preferably, the monomer stabilizer is at least one of reduced glutathione, DDT, glycine and nipagin Jin Bingku, the monomer stabilizer can reduce the formation of the complex by the monomers in the cleaning process, the concentration of the monomer stabilizer is 1-50mmol/L, a large number of experiments prove that the concentration of the monomer stabilizer is critical to the reduction of the formation of the complex by the monomers in the cleaning process, the surface charge or disulfide bond of the protein can be stabilized when one or more related reagents are in low concentration, but the protein denaturation or disulfide bond opening can be caused when the concentration is high, so that the possibility of the formation of the complex by the monomers can be avoided when the concentration and the proportion are proper, and the existence form of the complex can be maintained.

Preferably, the surfactant is a combination of nonionic and amphoteric surfactants, and the nonionic surfactant may include, but is not limited to, the following: tritonX-100, tween 20, tween 80, sodium lauryl sulfate, brij-35, sodium lauryl silicate, amphoteric surfactants can include, but are not limited to, the following: the CHAPS has the total concentration of the surfactant of 0.02-0.15% w/v, the surfactant can effectively remove nonspecific adsorption substances in the immune reaction process, a large number of experiments prove that the combination of the nonionic surfactant and the amphoteric surfactant can achieve the optimal washing effect, the specific binding of protein is not destroyed while the nonspecific adsorption substances are removed, and meanwhile, stable complexes formed between thrombin monomers and antithrombin complex monomers can be reduced, so that the detection result is more reliable and accurate.

Preferably, the cleaning solution further comprises a preservative, wherein the preservative is Proclin300 and/or Bronidox, the concentration of the preservative is 0.02% -1.2% w/v, the antibacterial effect of the cleaning solution is evaluated by detecting the blank of the cleaning solution through further screening and limiting the concentration of the preservative, if the antibacterial effect of the cleaning solution is poor, the blank luminous value of the cleaning solution is increased if the bacterial infection phenomenon occurs, the luminous value is increased more seriously, the bacterial infection is serious, and the jump value phenomenon occurs, and experiments prove that the cleaning solution can still keep the CV of the blank of the cleaning solution within 5% after being placed at 37 ℃ for 6 months, and the luminous value change of the cleaning solution is minimum compared with that of 37 ℃ for 6 months, so that the antibacterial effect of the cleaning solution is the best.

Preferably, the defoaming agent is a silicone oil type and/or silicone ether type defoaming agent, the concentration of the defoaming agent is 0.03-0.5% w/v, the CV of the cleaning solution added with the defoaming agent on a TAT project is smaller, and the signal to noise ratio is higher, so that after the defoaming agent is added into the cleaning solution, the generation of bubbles can be eliminated or inhibited, the mixing and cleaning of a reaction system are more sufficient, the occurrence of the jump value phenomenon in the detection process is effectively reduced, and the precision and the accuracy of reagent detection are improved.

A great number of experiments prove that the pH value of the cleaning solution is 7.1-7.5, and the pH value of the cleaning solution is between 7.1 and 7.5, so that a proper acid-base environment can be provided for the whole reaction system, the TAT protein captured by the antibody on the magnetic ball can not fall off, and the structure of the magnetic ball-antibody-compound TAT protein trimer can be well maintained.

A method for preparing a cleaning solution for thrombin-antithrombin complex (TAT) detection, comprising the steps of:

step one: inorganic salt, metal ions, buffering agent and surfactant are uniformly mixed and then defoamer is added;

step two: and (3) sequentially adding the targeted compound stabilizer and the monomer stabilizer into the mixed solution obtained in the step (A), and uniformly mixing to obtain the cleaning solution.

Advantageous effects

Compared with the prior art, the application provides a cleaning solution for thrombin-antithrombin complex (TAT) detection and a straw fermentation method, and has the following beneficial effects:

1. the stability of thrombin-antithrombin complex (TAT) in plasma can be improved by the targeted complex stabilizer in the cleaning liquid, complex dissociation is not easy to occur along with the cleaning, the possibility of forming a new complex by combining other salt ions, surfactants and saccharide components can be reduced, the combination of the surfactants and the targeted complex stabilizer is proved by a large number of realization, so that the TAT complex has better stability, bubbles generated in the process of adding the cleaning liquid or cleaning can be reduced by the defoamer, the occurrence of jumping value phenomenon can be reduced, and the detection accuracy and precision can be improved, so that the cleaning liquid can reduce non-specific interfering substances, reduce the degradation or denaturation of an object to be detected, and improve the accuracy and stability of a detection result;

2. the metal ions of the cleaning liquid can be provided by inorganic salts or other components containing the metal ions, the inorganic salts can be sodium chloride or potassium chloride, the sodium ions or the potassium ions are provided by sodium chloride or potassium chloride, the concentration of the metal ions is 100-500mM, the inorganic salts and the metal ions play a role in providing a function of maintaining the surface structure of protein, the cleaning liquid can have certain ionic strength, so that the antibodies can play a binding role in characteristics, and simultaneously thrombin-antithrombin complex (TAT) can exist stably;

3. the concentration of the monomer stabilizer of the cleaning solution is 1-50mmol/L, and a large number of experiments prove that the concentration of the monomer stabilizer is critical to reducing the effect of forming a compound by the monomer in the cleaning process;

4. the surfactant of the cleaning liquid is a nonionic surfactant and an amphoteric surfactant which are used in combination, the concentration of the surfactant is 0.02% -0.15% w/v, the surfactant can effectively remove nonspecific adsorption substances in the immune reaction process, a large number of experiments prove that the nonionic surfactant and the amphoteric surfactant are used in combination, the optimal cleaning effect can be achieved, the specific combination of proteins is not destroyed while the nonspecific adsorption substances are removed, meanwhile, stable complexes formed between thrombin monomers and antithrombin complex monomers can be reduced, and finally the detection result is more reliable and accurate;

5. the preservative of the cleaning solution is Proclin300 and/or nipagin Jin Bingku, the concentration of the preservative is 0.02% -1.2% w/v, the cleaning solution can still keep the blank CV of the cleaning solution within 5% after being placed at 37 ℃ for 6 months, and the change of the luminescence value is minimum compared with the blank CV of the cleaning solution at 37 ℃ for 6 months, so that the cleaning solution has the best antibacterial effect;

6. the defoaming agent of the cleaning fluid is a silicone oil type and/or silicone ether type defoaming agent, the concentration of the defoaming agent is 0.03-0.5% w/v, the CV of the cleaning fluid added with the defoaming agent on a TAT project is smaller, and the signal to noise ratio is higher, so that after the defoaming agent is added into the cleaning fluid, the generation of bubbles is eliminated or inhibited, the occurrence of value jump phenomenon in the detection process is effectively reduced, and the precision and accuracy of reagent detection are improved;

7. the pH value of the cleaning liquid is 7.1-7.5, and a large number of experiments prove that the pH value of the cleaning liquid is 7.1-7.5, so that a proper acid-base environment can be provided for the whole reaction system.

Detailed Description

The present application provides a wash solution for thrombin-antithrombin complex (TAT) detection, comprising: inorganic salts, metal ions, buffers, targeted complex stabilizers, monomer stabilizers, surfactants, preservatives, and defoamers.

The metal ions can be provided by the inorganic salt, the inorganic salt can be sodium chloride or potassium chloride, the sodium ions or potassium ions are provided by sodium chloride or potassium chloride, the metal ions can also be added by other components containing the metal ions, and the concentration of the metal ions is 100-500mM.

The pH value of the buffer is 7.0-7.5, the concentration of the buffer is 10-100mM, the buffer adopts Tris buffer, and the pH value of the buffer is regulated to 7.0-7.5 by adopting a solid acid regulator before use.

The targeted compound stabilizer is at least one of mannitol, trehalose and arginine, and the concentration of the targeted compound stabilizer is 0.1% -0.5% w/v.

The monomer stabilizer is at least one of reduced glutathione, DDT, glycine and nipagin Jin Bingku, the concentration of the monomer stabilizer is 1-50mmol/L, and a large number of experiments prove that the concentration of the monomer stabilizer is critical to reducing the effect of forming a compound by a monomer in the cleaning process of the compound.

The surfactant is a nonionic surfactant and an amphoteric surfactant which are used in a combined way, the concentration of the surfactant is 0.02% -0.15% w/v, and a large number of experiments prove that the nonionic surfactant and the amphoteric surfactant are used in a combined way, so that the optimal washing effect can be achieved, the specific combination of proteins is not destroyed while the non-specific adsorption substances are removed, and meanwhile, a stable compound can be formed between thrombin monomers and antithrombin compound monomers, and finally, the detection result is more reliable and accurate.

The defoaming agent is a silicone oil type and/or silyl ether type defoaming agent, the concentration of the defoaming agent is 0.03-0.5% w/v, the CV of the cleaning fluid added with the defoaming agent on a TAT project is smaller, and the signal to noise ratio is higher, so that after the defoaming agent is added into the cleaning fluid, the generation of bubbles is eliminated or inhibited, the occurrence of the jump value phenomenon in the detection process is effectively reduced, and the precision and the accuracy of reagent detection are improved.

A large number of experiments prove that the pH value of the cleaning solution is 7.1-7.5, and the pH value of the cleaning solution is 7.1-7.5, so that a proper acid-base environment can be provided for the whole reaction system.

It should be noted that the cleaning solution further comprises a preservative, the preservative is Proclin300 and/or Bronidox, the concentration of the preservative is 0.02% -1.2% w/v, the cleaning solution blank is detected to evaluate the antibacterial effect of the cleaning solution by further screening and limiting the concentration of the preservative, if the antibacterial effect of the cleaning solution is poor, the blank luminous value of the cleaning solution is increased if the cleaning solution has a bacterial infection phenomenon, the luminous value is increased more seriously, the bacterial infection is serious, and the jump value phenomenon appears, and experiments prove that the cleaning solution can still keep the CV of the blank of the cleaning solution within 5% after being placed at 37 ℃ for 6 months, and the luminous value change of the cleaning solution is minimum compared with that of the cleaning solution at 37 ℃ for 6 months, so that the antibacterial effect of the cleaning solution is the best.

A method for preparing a cleaning solution for thrombin-antithrombin complex (TAT) detection, comprising the steps of:

step one: adjusting the pH value of the buffer solution to 7.0-7.5 by using a solid acid regulator, sequentially adding inorganic salt, metal ions, a buffering agent and a surfactant, uniformly mixing, adding a defoaming agent, and then carrying out constant volume by using a solvent;

step two: and (3) sequentially adding the targeted compound stabilizer and the monomer stabilizer into the mixed solution obtained in the step (A), and uniformly mixing to obtain the cleaning solution.

The solid acid modulator used in step one comprises sulfamic acid and/or boric acid.

26 example cleaning solutions and 3 comparative example cleaning solutions with different proportions are used, the cleaning solutions are prepared and diluted according to the proportions, and then the 26 example cleaning solutions and the 3 comparative example cleaning solutions are measured by using a Shine 1000 full-automatic chemiluminescence immunoassay of Shenzhen YingKai biotechnology Co., ltd, and the results are shown in the following table:

the samples described in the table are thrombin-antithrombin III complex (TAT)

/>

/>

/>

/>

/>

/>

/>

/>

/>

/>

/>

/>

/>

As shown in examples one to three in the table, only the pH value of the cleaning solution in examples one to three is different, wherein the pH value of the cleaning solution in example two is 7.3, and the CV value of the measurement result is 1.

As shown in examples four to six in the table, the concentration of sodium ion is only different in examples four to six, wherein the concentration of sodium ion in the cleaning solution of example four is 230mM, and the CV value is 3, and compared with the most preferred examples five and six, the metal ion functions to maintain the surface structure of the protein, and the appropriate concentration of the metal ion in the system can provide the cleaning solution with appropriate ionic strength, thereby allowing the antibody to perform the binding function characteristically, and also allowing the thrombin-antithrombin complex (TAT) to exist stably.

As compared with example four and example seven in the table, the targeted complex stabilizer of example four is mannitol, the targeted complex stabilizer of example seven is mannitol and arginine which are commonly used, the concentrations of the targeted complex stabilizers of 2 examples are consistent, the CV value of implementation four is 3, and the CV value of implementation seven is 2; as compared with example five and example eight in the table, the targeted complex stabilizer of example five is mannitol, the targeted complex stabilizer of example eight is mannitol and arginine, the concentrations of the 2 targeted complex stabilizers of examples are consistent, the CV value of the implementation five is 4, and the CV value of the implementation eight is 3.5; as compared with example six and example nine in the table, the targeted complex stabilizer of example six is mannitol, the targeted complex stabilizer of example nine is mannitol and arginine, the concentrations of the 2 targeted complex stabilizers of examples are consistent, the CV value of the implementation six is 3.5, and the CV value of the implementation nine is 3; the comparison shows that the mannitol and the arginine are used together at the same concentration as compared with single mannitol, the CV value of the measurement result is better, and the CV value of the measurement result is better than that of one of the two targeted complex stabilizers.

As shown in examples ten to twelve in the table, the concentrations of reduced glutathione were different from each other in examples ten to twelve, wherein the concentration of reduced glutathione in example eleven was 30mmol/L and the CV value was 3, and the measurement results were better than those in examples ten and twelve, since the low concentration of the monomer stabilizer was capable of stabilizing the surface charge or disulfide bond of the protein, but the high concentration resulted in denaturation of the protein or opening of disulfide bond, the proper concentration and ratio of the monomer stabilizer was capable of avoiding the possibility of forming a complex of the monomer and maintaining the existence form of the complex.

As compared with example thirteenth and example tenth in the table, the monomer stabilizer of example tenth is reduced glutathione, the monomer stabilizer of example thirteenth is both reduced glutathione and glycine, the concentrations of the monomer stabilizers of 2 examples are identical, the CV value of implementation ten is 4, and the CV value of implementation thirteen is 3.5; as compared with example fourteen and example eleventh in the table, the monomer stabilizer of example eleventh is reduced glutathione, the monomer stabilizer of example fourteen is used together with both reduced glutathione and glycine, the concentrations of the monomer stabilizers of 2 examples are identical, the CV value of implementation fourteen is 2, and the CV value of implementation eleven is 3; as compared with the fifteen and twelve examples in the table, the monomer stabilizer of the twelve examples is reduced glutathione, the monomer stabilizer of the fifteen examples is used together with the two types of reduced glutathione and glycine, the concentrations of the 2 example monomer stabilizers are consistent, the CV value of fifteen implementation is 4, and the CV value of twelve implementation is 5; the comparison shows that the single concentration of the monomer stabilizer, the reduced glutathione and the reduced glutathione are used together to obtain better CV value of the measurement result compared with the single concentration of the reduced glutathione, and the comparison shows that the single concentration of the monomer stabilizer is used together to obtain better CV value of the measurement result compared with the single concentration of the monomer stabilizer.

As shown in the table, the surfactant of the thirteenth example is CHAPS and Triton X-100, the nonionic surfactant and the amphoteric surfactant are used together, the surfactant of the thirteenth example is Triton X-100, the concentration of the surfactant of the 2 examples is consistent, the CV value of the implemented thirteen is 3.5, and the CV value of the implemented sixteen is 3; as shown in the table, the surfactant of the seventeenth example was CHAPS and Triton X-100, the nonionic surfactant and the amphoteric surfactant were used in combination, the surfactant of the fourteen example was Triton X-100, the concentration of the surfactant of the 2 examples was uniform, the CV value of the fourteen examples was 2, and the CV value of the seventeen examples was 1.5; as shown in the table, compared with the eighteen examples, the eighteen examples are used together with CHAPS and Triton X-100, the nonionic surfactant and the amphoteric surfactant are used together, the fifteen examples are used together with Triton X-100 as a single surfactant, the concentration of the 2 examples is consistent, the CV value of fifteen examples is 4, and the CV value of eighteen examples is 3.5; the comparison shows that the combination of the surfactant with the same concentration, the nonionic surfactant and the amphoteric surfactant has better CV value than the single surfactant.

As shown in the examples nineteenth to twenty-first embodiments, only the concentration of the silicone oil type defoamer is different in the examples nineteenth to twenty-first embodiments, wherein the concentration of the silicone oil type defoamer in the example twenty is 0.3% w/v, and the CV value of the measurement result is 2.

The two defoamers of the twenty-second embodiment are silicone oil defoamers and silicone ether defoamers and are used together, the two defoamers of the twenty-first embodiment are silicone oil defoamers, the concentration of the 2 embodiment surfactants is the same, the CV value of the twenty-second embodiment is 1, the CV value of the twenty-second embodiment is 2, the three defoamers of the twenty-third embodiment are silicone oil defoamers and silicone ether defoamers and are used together, the one defoamer of the twenty-first embodiment is silicone oil defoamers, the concentration of the 2 embodiment surfactants is the same, the CV value of the one embodiment is 2.5, the one defoamers of the silicone oil defoamers and the silicone ether defoamers are used together, the CV value of the two defoamers of the comparison finding is better, and the CV value of the two defoamers is better than that of one defoamer.

As shown in examples twenty-four to twenty-six in the table, only Tris-salt buffer concentrations were different in examples twenty-four to twenty-five, wherein Tris-salt buffer concentration of example twenty-six was 100mM, and CV value was 2, and the measurement results were better than those of examples twenty-four and twenty-five.

As shown in the first example and the first comparative example, the cleaning solution of the first comparative example and the first comparative example do not contain silicone oil type defoamer, and under the condition that other conditions are unchanged, the CV value measured by the first comparative example is 10, the CV value measured by the first example is 3, and the CV value measured by the first comparative example is better.

As shown in the second and the second comparative examples, the pH value of the cleaning solution of the second comparative example is 8, the pH value of the cleaning solution of the second comparative example is 7.3, the CV value measured in the first comparative example is 9, the CV value measured in the first example is 1, the CV value measured in the first example is better, and a great amount of experiments prove that the pH value of the cleaning solution is 7.3, which can provide a proper acid-base environment for the whole reaction system, can prevent the TAT protein captured by the antibody on the magnetic ball from falling off, and better maintain the structure of the TAT protein trimer of the magnetic ball-antibody-complex.

As shown in the table of the third embodiment and the third comparative embodiment, the cleaning solution of the third comparative embodiment and the third comparative embodiment does not contain the targeted stable complex stabilizer, and under the condition that other conditions are not changed, the CV value measured by the third comparative embodiment is 10, the CV value measured by the third embodiment is 2.5, the CV value measured by the third embodiment is better, the targeted complex stabilizer plays a role in stabilizing the complex, and the phenomenon of complex dissociation caused by cleaning can be reduced, so that the CV value measured by the targeted complex stabilizer is better.

Claims (10)

1. A wash solution for thrombin-antithrombin complex (TAT) detection, comprising: inorganic salts, metal ions, buffers, targeted complex stabilizers, monomer stabilizers, surfactants, and defoamers.

2. A cleaning solution for thrombin-antithrombin complex (TAT) detection according to claim 1, wherein the metal ions are provided by the inorganic salt.

3. A cleaning solution for thrombin-antithrombin complex (TAT) detection according to claim 1, wherein the buffer has a pH of 7.0 to 7.5.

4. The wash solution for thrombin-antithrombin complex (TAT) detection according to claim 1, wherein the targeted complex stabilizer is at least one of mannitol, trehalose, arginine.

5. The wash solution for thrombin-antithrombin complex (TAT) detection according to claim 1, wherein the monomer stabilizer is at least one of reduced glutathione, DDT, glycine, nipagin Jin Bingku.

6. A cleaning solution for thrombin-antithrombin complex (TAT) assay according to claim 1, wherein the surfactant is a combination of a nonionic surfactant and an amphoteric surfactant.

7. The wash solution for thrombin-antithrombin complex (TAT) detection according to claim 1, wherein the wash solution further comprises a preservative, which is Proclin300 and/or Bronidox.

8. The washing liquid for thrombin-antithrombin complex (TAT) detection according to claim 1, wherein the antifoaming agent is of silicone oil type and/or of silicone ether type.

9. A wash solution for thrombin-antithrombin complex (TAT) assay according to any one of claims 1 to 8, wherein the pH of the wash solution is from 7.1 to 7.5.

10. A method for preparing a cleaning solution for thrombin-antithrombin complex (TAT) detection, comprising the steps of:

step one: inorganic salt, metal ions, buffering agent and surfactant are uniformly mixed and then defoamer is added;

step two: and (3) sequentially adding the targeted compound stabilizer and the monomer stabilizer into the mixed solution obtained in the step (A), and uniformly mixing to obtain the cleaning solution.