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Preparation method of polystyrene microspheres of coupled antibodies

A polystyrene microsphere and antibody technology, applied in the preparation of microspheres, microcapsule preparations, etc., can solve the problems of easy detachment and limitation of antibodies, and achieve the effects of reducing process difficulty, reducing process cost, and simplifying operation steps.

Active Publication Date: 2019-04-16
蓝怡(湖南)医疗器械有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This method uses tangential flow filtration technology to separate the antibody-coupled latex microspheres from the residual antibody. Although it overcomes the shortcomings of easy shedding and inactivation of the antibody in the centrifugation process, it is still difficult to achieve in mass production. Restricted

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0052] This example provides a method for preparing antibody-coupled carboxypolystyrene microspheres:

[0053] (1) Carboxylated polystyrene microspheres are placed in an MES buffer solution with a concentration of 50 mM and a pH value of 5.7, and the concentration of diluted carboxypolystyrene microspheres is adjusted to 0.25%;

[0054] (2) Add the antibody to the buffer in step (1), adjust the final concentration of the antibody to 0.7 mg / mL, add 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, Coupling reaction at pH 5.7 for 12 hours;

[0055] (3) ultrasonically mix the solution after the reaction in step (2), and add BSA to make the final concentration of BSA reach 2%;

[0056] (4) Add surfactant sodium lauryl sulfate to the solution after step (3) sealing, make sodium lauryl sulfate final concentration be 0.3%, stir and mix 6h at 25 ℃, add the MOPSO of 50mM and The mixed buffer composed of Tris was diluted to the use concentration, adjusted to pH 8.0, and sto...

Embodiment 2

[0058] This example provides a method for preparing antibody-coupled carboxypolystyrene microspheres:

[0059] (1) Carboxylated polystyrene microspheres are placed in an MES buffer solution with a concentration of 40 mM and a pH value of 5.5, and the concentration of diluted carboxypolystyrene microspheres is adjusted to 0.25%;

[0060] (2) Add the antibody to the buffer in step (1), adjust the final concentration of the antibody to 1 mg / mL, add 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, and Coupling reaction at pH 5.5 for 24 hours;

[0061] (3) ultrasonically mix the solution after the reaction in step (2), and add BSA to make the final concentration of BSA reach 1.5%;

[0062] (4) Add the surfactant sodium dodecylbenzene sulfonate to the solution after step (3) is closed, so that the final concentration of sodium dodecylbenzene sulfonate is 0.5%, stir and mix at 18°C ​​for 4h, add The mixed buffer composed of 50 mM MOPSO and Tris was diluted to the use co...

Embodiment 3

[0064] This example provides a method for preparing antibody-coupled carboxypolystyrene microspheres:

[0065] (1) Carboxylated polystyrene microspheres are placed in an MES buffer solution with a concentration of 60 mM and a pH value of 6, and the concentration of diluted carboxypolystyrene microspheres is adjusted to 0.3%;

[0066] (2) Add the antibody to the buffer in step (1), adjust the final concentration of the antibody to 0.5 mg / mL, add 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, Coupling reaction at pH 6 for 1 h;

[0067] (3) Mix the solution after step (2) reaction, add BSA, make the final concentration of BSA reach 2%;

[0068] (4) add surfactant sodium dodecylsulfate and sodium dodecylbenzenesulfonate to the solution after step (3) sealing, make sodium dodecylsulfate and sodium dodecylbenzenesulfonate total The final concentration of the solution was 0.5%, mixed at 25°C for 4 hours, diluted to the use concentration by adding a mixed buffer compos...

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Abstract

The invention provides a preparation method of polystyrene microspheres of coupled antibodies. The preparation method comprises the following steps of (1) placing carboxyl polystyrene microspheres ina buffer solution and adjusting the concentration of the carboxyl polystyrene microspheres; (2) adding the antibodies into the buffer solution in step (1) and adding a coupling agent for a coupling reaction; (3) uniformly mixing the solution obtained after the reaction in step (2) and adding a sealing reagent; (4) adding a surfactant into the sealed solution in step (3) and uniformly blending themixed solution to obtain the polystyrene microspheres of the coupled antibodies. According to the provided preparation method, the centrifugation and ultrafiltration processes are omitted, the processdifficulty is lowered, the production cost is reduced at the same time, and the preparation method has a high value in practical application and production.

Description

technical field [0001] The invention belongs to the field of biological materials and relates to a preparation method of polystyrene microspheres coupled with antibodies. Background technique [0002] There are two main methods for coupling microspheres and antibodies: one is to use chemical coupling agents to generate covalent bonds between functionalization (carboxyl / amino / aldehyde groups, etc.) and antibodies, and the other is to The physical adsorption method relies on electrostatic forces to adsorb antibodies to the surface of microspheres. In order to remove unbound antibodies, both of these methods involve the steps of filtration and washing. In the process of filtration and washing, centrifugation and ultrafiltration are generally used. However, these cleaning and filtration processes are time-consuming, and the batch size of production is limited by centrifuges or ultrafiltration equipment, which increases the cost of finished products. [0003] CN104530459A discl...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): B01J13/02
CPCB01J13/02
Inventor 李子樵王森
Owner 蓝怡(湖南)医疗器械有限公司
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