Method for coupling carboxyl microspheres with amino groups, sensitizing latex and kit

A microsphere and coupling technology, which is applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of limited reaction conditions, cumbersome operation steps, harsh operating conditions, etc., to achieve simple operation, overcome cumbersome operation, and high-efficiency coupling Effect

Pending Publication Date: 2020-08-25
JILIN GETEIN BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Therefore, both the one-step method and the two-step method in the prior art use EDC as a crosslinking agent, resulting in many restrictions on the reaction conditions. The harsh operating conditions and the cumbersome operation steps have further led to the current method of coupling amino groups with carboxyl microspheres. High cost and low coupling efficiency
[0006] N,N'-Carbonyldiimidazole (CDI) is a reagent for activating carboxylic acids in solution or solid-phase peptide synthesis, but it has not been used in the field of carboxyl microsphere labeling proteins so far

Method used

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  • Method for coupling carboxyl microspheres with amino groups, sensitizing latex and kit
  • Method for coupling carboxyl microspheres with amino groups, sensitizing latex and kit
  • Method for coupling carboxyl microspheres with amino groups, sensitizing latex and kit

Examples

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Effect test

Embodiment 1

[0042] According to the antistreptolysin "O" test kit prepared by the preferred scheme of the present invention, its steps are:

[0043](1) Preparation: prepare polystyrene latex microspheres of 155nm, and dilute the latex microspheres with 50mM / L 2-(N-morpholine)ethanesulfonic acid at pH 6.0 to a final concentration of 1.5% w / v to obtain latex microspheres. Ball solution; N,N'-carbonyldiimidazole (CDI) was dissolved with 50mM / L 2-(N-morpholine)ethanesulfonic acid at pH 6.0 to obtain a CDI solution with a concentration of 0.01g / mL;

[0044] (2) Activation: Based on the weight of the latex microspheres, add each mg of microspheres dropwise to the latex microsphere solution in the proportion of 50 μL of CDI solution, shake and mix, shake and activate at 25°C for 25 minutes, and the shaking speed is 200 rpm / min. Obtain activated latex microspheres;

[0045] (3) Add 50mM / L phosphate buffer to adjust the pH to 8.0;

[0046] (4) Coupling: based on the weight of the latex microsphe...

Embodiment 2

[0050] According to the C-reactive protein kit prepared by the preferred scheme of the present invention, its steps are:

[0051] (1) Preparation: Prepare 107nm polystyrene latex microspheres, and the latex microspheres are diluted with 50mM / L 2-(N-morpholine)ethanesulfonic acid at pH 6.0 to a final concentration of 3% w / v to obtain latex microspheres. Ball solution; N,N'-carbonyldiimidazole (CDI) was dissolved with 50mM / L 2-(N-morpholine)ethanesulfonic acid at pH 6.0 to obtain a CDI solution with a concentration of 0.01g / mL;

[0052] (2) Activation: Based on the weight of the latex microspheres, add each mg of microspheres dropwise to the latex microsphere solution in the proportion of 80 μL of CDI solution, shake and mix, shake and activate at 30 ° C for 30 minutes, and the shaking speed is 100 rpm / min. Obtain activated latex microspheres;

[0053] (3) Add 50mM / L borate buffer to adjust the pH to 7.5;

[0054] (4) Coupling: based on the weight of the latex microspheres, ad...

Embodiment 3

[0059] According to the β2 microglobulin test kit prepared by the preferred scheme of the present invention, its steps are:

[0060] (1) Preparation: Prepare 123nm polystyrene latex microspheres, which are diluted with 50mM / L 2-(N-morpholine)ethanesulfonic acid at pH 6.0 to a final concentration of 1.8% w / v to obtain latex microspheres. Ball solution; N,N'-carbonyldiimidazole (CDI) was dissolved with 50mM / L 2-(N-morpholine)ethanesulfonic acid at pH 6.0 to obtain a CDI solution with a concentration of 0.01g / mL;

[0061] (2) Activation: Based on the weight of the latex microspheres, add each mg of microspheres dropwise to the latex microsphere solution in the proportion of 60 μL of CDI solution, shake and mix, shake and activate at 37°C for 20 minutes, and the shaking speed is 200 rpm / min. Obtain activated latex microspheres;

[0062] (3) Add 50mM / L borate buffer to adjust the pH to 7.5;

[0063] (4) Coupling: based on the weight of the latex microspheres, add the polyclonal a...

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Abstract

The invention provides a method for coupling carboxyl microspheres with amino, sensitizing latex and a kit, and belongs to the technical field of medical reagents. The method comprises the following steps: dropwise adding a CDI solution into a latex microsphere solution, and oscillating and uniformly mixing to obtain activated latex microspheres; adding a coupling buffer solution into the activated latex microspheres to adjust the pH value; adding an antibody into the solution of which the pH value is adjusted, and oscillating and uniformly mixing to obtain a coupled solution; and adding the bovine serum albumin mother liquor into the coupled solution, and oscillating and uniformly mixing to obtain the sensitizing latex. The invention also provides the sensitizing latex prepared by the preparation method. The invention also provides a kit containing the sensitizing latex. The method is high in coupling efficiency and low in cost, and the obtained kit is excellent in performance.

Description

technical field [0001] The invention belongs to the technical field of medical reagents, and in particular relates to a method for coupling amino groups with carboxyl microspheres, sensitizing latex and a kit. Background technique [0002] The main methods of in vitro diagnosis are enzyme-linked immunoassay, latex turbidimetry, chemiluminescence and molecular diagnosis. Latex nephelometry and chemiluminescence methods involve the use of microsphere-labeled proteins. The method of microsphere labeling protein (antigen or antibody or avidin) is mainly adsorption method and covalent coupling method, the purpose is to coat the antibody or antigen corresponding to the substance to be tested on the microsphere, through The sensitivity of the test is improved by detecting the characteristics of the reacted microspheres coated with the corresponding antigen and antibody. The surface of microspheres commonly used in the covalent coupling method is coupled with functional groups - c...

Claims

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Application Information

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IPC IPC(8): G01N33/531
CPCG01N33/531
Inventor 陈凯丽刘玲徐李鹏
Owner JILIN GETEIN BIOTECH CO LTD
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