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Preparation method and application of D-dimer immuno latex microspheres

A technology of dimer and polystyrene microspheres, which is applied in the field of medical immunodiagnosis, can solve the problems of low product yield, coupling efficiency, and antibody activity, and achieve high sensitivity, good clinical application prospects, and accurate quantification. Effect

Active Publication Date: 2017-01-04
WUHAN KING DIAGNOSTIC TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] The purpose of the present invention is to overcome the defect of low product yield in the existing D-dimer immune latex microsphere preparation method, to provide a D-dimer immune latex with high coupling efficiency and little influence on antibody activity after coupling Microsphere preparation method and D-dimer determination kit by latex-enhanced immune turbidimetric method using the D-dimer immune latex microspheres

Method used

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  • Preparation method and application of D-dimer immuno latex microspheres

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Effect test

Embodiment 1

[0054] The preparation method of D-dimer immune latex microspheres provided by the present invention is as follows:

[0055] 1. Prepare polystyrene microspheres with a particle size of 240nm, the concentration is 10%, and take 2ml; the polystyrene microspheres are first diluted with 50mmol / L 2-morpholineethanesulfonic acid buffer to a final concentration of 0.5% w / v, to obtain a microsphere solution with a pH of 5.5 to 6.5, and the volume of the solution is 40 mL;

[0056] 2. Dissolve carbodiimide in 2-morpholineethanesulfonic acid buffer to obtain a carbodiimide solution with a concentration of 10 mg / mL;

[0057] 3. Based on the weight of the polystyrene microspheres contained in the microsphere solution, add the carbodiimide solution to the microsphere solution at a rate of 35~45μL per 10mg polystyrene microspheres. 80μL, shaking and activating at 4℃ for 1h, shaking speed of 50r / min, to obtain activated polystyrene microsphere solution;

[0058] 4. Dissolve the DD monoclonal antibo...

Embodiment 2

[0061] The preparation method of D-dimer immune latex microspheres provided by the present invention is as follows:

[0062] 1. Prepare polystyrene microspheres with a particle size of 200nm, the concentration is 10%, and take 2ml; the polystyrene microspheres are first diluted with a concentration of 50mmol / L 2-morpholineethanesulfonic acid buffer to a final concentration of 0.4% w / v, to obtain a microsphere solution with a pH of 5.5 to 6.5, and the volume of the solution is 50 mL;

[0063] 2. Dissolve carbodiimide in 2-morpholineethanesulfonic acid buffer to obtain a carbodiimide solution with a concentration of 10 mg / mL;

[0064] 3. Based on the weight of the polystyrene microspheres contained in the microsphere solution, add the carbodiimide solution to the microsphere solution at a rate of 35~45μL per 10mg polystyrene microspheres. 80μL, shaking and activating at 8℃ for 1h, shaking speed of 30r / min, to obtain activated polystyrene microsphere solution;

[0065] 4. Dissolve the D...

Embodiment 3

[0068] The preparation method of D-dimer immune latex microspheres provided by the present invention is as follows:

[0069] 1. Prepare polystyrene microspheres with a particle size of 400nm, the concentration is 10%, and take 2ml; the polystyrene microspheres are first diluted with a concentration of 50mmol / L 2-morpholineethanesulfonic acid buffer to a final concentration of 0.6% w / v, to obtain a microsphere solution with a pH of 5.5 to 6.5, and the volume of the solution is 40 mL;

[0070] 2. Dissolve carbodiimide in 2-morpholineethanesulfonic acid buffer to obtain a carbodiimide solution with a concentration of 10 mg / mL;

[0071] 3. Based on the weight of the polystyrene microspheres contained in the microsphere solution, add the carbodiimide solution to the microsphere solution at a rate of 35~45μL per 10mg polystyrene microspheres. 84μL, shaking and activating at 2℃ for 1h, shaking speed of 50r / min, to obtain activated polystyrene microsphere solution;

[0072] 4. Dissolve the D...

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Abstract

The invention discloses a preparation method and application of D-dimer immuno latex microspheres. In the preparation method of D-dimer immuno latex microspheres, a chemical crosslinking method is optimized, the carboxyl of polystyrene microsphere is activated in a low-pH condition and then coupled with amino of an antibody dissolved in a high-pH condition, and the mixture of the two is neutral and is coupled for 12-24h at a low temperature, so that not only can the coupling efficiency of the antibody be guaranteed, but also the damage of the antibody activity caused by an activator is reduced to a great extent. A latex enhanced turbidimetric immunoassay D-dimer test kit prepared from the immuno latex microspheres prepared by the method has the characteristics of high sensitivity, accurate quantification, good repeatability and stable property, and the lowest detection limit can reach 0.01mg / L; the latex enhanced turbidimetric immunoassay D-dimer test kit can be applied to a full-automatic biochemical analyzer or coagulation analyzer, is quick and easy to operate, only needs 5-10 minutes from detection to result collection and has relatively good clinical application prospect.

Description

Technical field [0001] The invention relates to the field of medical immunodiagnosis, in particular to a preparation method and application of D-dimer immune latex microspheres. Background technique [0002] D-dimer is a specific degradation product produced by fibrin monomer cross-linked by activating factor XIII and then hydrolyzed by plasmin. It is a specific marker of fibrinolysis process. [0003] The measurement of plasma D dimer is an experiment to understand the function of secondary fibrinolysis. The principle of the latex-enhanced immunoturbidimetric method used in the prior art is: anti-DD monoclonal antibody is coated on latex particles, if D-dimer exists in the recipient plasma, an antigen-antibody reaction will occur, and the latex particles will aggregate phenomenon. [0004] In 1977, Sternberg created the rate-scattering turbidimetric method to determine the amount of antigen in a sample by measuring the degree of scattering of light by a solution. Light of a certa...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/543
CPCG01N33/54346
Inventor 王明李华中吴艳芳
Owner WUHAN KING DIAGNOSTIC TECH CO LTD
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