Kit for detecting alpha 1-microglobulin and preparation method of kit

A technology of microglobulin and kit, which is applied in the direction of biological testing, measuring devices, material inspection products, etc., can solve the problems of poor stability of physical adsorption method, falling off of antibodies, and degradation of reagent performance, so as to reduce production and preparation costs, reduce The effect of high usage and high accuracy

Inactive Publication Date: 2017-07-07
SHANGHAI FOSUN LONG MARCH MEDICAL SCI CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The preparation process generally includes physical adsorption method and chemical coupling method; among them, the physical adsorption method has poor stability, and the antibody is easy to fall off from the latex particles, resulting in a decrease in the performance of the reagent; while the chemical coupling method uses a functional Group (such as carboxyl) latex microspheres, under the action of an activator such as EDC (1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride), can be conjugated to the antibody , antibody bound to microspheres
However, the chemical coupling method detects α 1 -MG's kit has strict requirements on the reaction system

Method used

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  • Kit for detecting alpha 1-microglobulin and preparation method of kit
  • Kit for detecting alpha 1-microglobulin and preparation method of kit
  • Kit for detecting alpha 1-microglobulin and preparation method of kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] Example 1: Detection of α 1 - Preparation of Microglobulin Kit

[0054] The kit of embodiment involves the main raw material of reagent as follows:

[0055] 1. Anti-human alpha 1 - Microglobulin Antibody (Shenzhen Feipeng Bio)

[0056] 2. Latex microspheres: use polystyrene latex particles with diameters of 68nm and 220nm with carboxyl groups (Jisha, Japan)

[0057] Prepare reagent R1: 50mM MOPS buffer, add 10g PEG6000, 1gNaN 3 , 1.5g BSA, 16gNaCl, 3g Tween-20, the rest is purified water, the total preparation volume is 1L, pH value is 8.0, the reagent is a colorless or light yellow transparent solution.

[0058] Reagent R2 formulation: 50mM MOPS buffer with 0.35% α 1 - Microglobulin antibody coated latex microspheres, 1.5% BSA, 1.7% NaCl, 1.3% sucrose, 0.08% NaN 3 , pH 7.7, the reagent is a milky white solution.

[0059] The steps for preparing reagent R2 are as follows:

[0060] (1) 1.75g ​​of 68nm latex microspheres (small particle size), add 10ml of MES solu...

Embodiment 2

[0081] Example 2: Detection of α 1 - Preparation of Microglobulin Kit

[0082] 1. the kit of the present invention relates to the main raw material of reagent as follows:

[0083] anti human alpha 1 - Microglobulin Antibody (Shenzhen Feipeng Bio)

[0084] Latex microspheres: use polystyrene latex particles with diameters of 95nm and 188nm with carboxyl groups (Japan Jie Shi Ya)

[0085] 2. the preparation of the main reagent of the present embodiment is as follows:

[0086] Reagent R1: 50mMH 3 BO 3 Buffer, add 0.4% PEG6000, 0.05% NaN 3 , 0.3% BSA, 0.8% NaCl, 0.1% Tween-20, pH 8.5, total preparation volume 1L, the reagent is a colorless or light yellow transparent solution.

[0087] Reagent R2: 50mMH 3 BO 3 Buffer with 0.15% alpha 1 - Microglobulin antibody coated latex microspheres, 0.4% BSA, 0.5% NaCl, 0.3% sucrose, 0.05% NaN 3 , pH 8.5, the reagent is a milky white solution. Concrete preparation steps are as follows:

[0088] (1) Add 1.2g of 68nm latex microsphe...

Embodiment 3

[0114] Example 3: α 1 - Microglobulin Reagent R2 Pilot Scale-Up (Tangential Flow Filtration)

[0115] During the scale-up process of R2 reagents from small samples to pilot plants, two methods, centrifugation and tangential flow filtration, are usually included; the centrifugation method is simple and easy to implement, but the amount of single preparation is small, and the difference between batches is difficult to control; in contrast, tangential flow filtration Filtration can better solve the problem of batch-to-batch variation. As long as the indicators are properly controlled, a large amount of R2 reagent can be prepared stably.

[0116] For the raw materials and reagent components used in this example, see Example 1 for details, the preparation steps are consistent with Example 1, and the total amount of R2 reagent prepared is 4L (68nm latex microsphere solution coupled to anti-α1-microglobulin antibody 2L ; 188nm latex microsphere solution 2L coupled with anti-α1-micro...

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Abstract

The invention discloses a kit for detecting alpha 1-microglobulin and a preparation method of the kit. The kit consists of a reagent R1 and a reagent R2 according to the volume ratio of 1: 1. The reagent R1 comprises 120-150 mmol/L of a buffer solution, 3-20 g/L of a stabilizer, 5-20 g/L of electrolyte, 3-20 g/L of a sensitizer, 0.5-10 g/L of a surfactant, 0.2-1 g/L of a preservative and purified water. The reagent R2 comprises 1.5-3.5 g/L of latex microspheres with two grain sizes, 20-150 mmol/L of a suffer solution, 3-20 g/L of a stabilizer, 5-20 g/L of electrolyte, 0.5 g/L of a preservative and purified water, wherein the latex microspheres with two grain sizes are respectively coupled with the same anti-human alpha 1-microglobulin polyclonal antibody. The pH value of the kit is 7.0-9.5. The kit is rapid in detection and high in accuracy. The production cost is greatly reduced. The kit is suitable for industrial production and has high application value.

Description

technical field [0001] The invention relates to biological reagents, in particular to a detection kit, in particular to a latex-enhanced immune turbidimetric method for detecting α 1 - Kits for microglobulins and methods for their preparation. Background technique [0002] alpha 1 - Microglobulin (α 1 -MG) is a glycoprotein widely distributed on the surface of body fluids and lymphocyte membranes. It consists of 167 amino acids and has a molecular weight of about 27kDa. It is mainly synthesized in the liver and lymphocytes; in blood, α 1 - Microglobulin exists in two forms: free α 1 - MG and alpha bound to IgA (immunoglobulin A) 1 -MG-1gA; free α 1 -MG can be freely filtered through the glomerulus, about 95%-99% is reabsorbed and metabolized in the renal tubules, and only a small amount is excreted in the urine; bound α 1 -MG-IgA cannot pass through the glomeruli, and its concentration in the urine is zero. alpha 1 -MG production in the body is constant and less affe...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68G01N33/543G01N33/545
CPCG01N33/68G01N33/54313G01N33/545G01N2333/47
Inventor 栾春杰景盛
Owner SHANGHAI FOSUN LONG MARCH MEDICAL SCI CO LTD
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