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Preparation method of anti-platelet antibody detection signal preparation, anti-platelet antibody detection signal preparation and application

An antibody detection and anti-platelet technology, which is applied in measuring devices, biological testing, material inspection products, etc., can solve problems such as complex reaction systems, safety risks, and short validity periods, and achieve simplified detection steps, extended storage time, and simplified composition effect

Pending Publication Date: 2021-04-16
宝锐生物科技泰州有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0009] The object of the present invention is to provide a preparation method of anti-platelet antibody detection signal preparation, and the detection signal preparation obtained by the method is used to replace the red blood cell preparation used in the existing platelet antibody detection technology, so as to alleviate the problems existing in the prior art. Technical issues such as short validity period, complex response system and safety risks

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  • Preparation method of anti-platelet antibody detection signal preparation, anti-platelet antibody detection signal preparation and application
  • Preparation method of anti-platelet antibody detection signal preparation, anti-platelet antibody detection signal preparation and application
  • Preparation method of anti-platelet antibody detection signal preparation, anti-platelet antibody detection signal preparation and application

Examples

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Embodiment 1

[0058] This example provides a method for purifying human IgG antibodies from human plasma, including rapidly thawing 1L of frozen human plasma stored in ACD preservation solution at 37°C, centrifuging at 8000g and 4°C for 20min, and then discarding the supernatant to obtain Component A. Component A was diluted with an equal volume of PBS and filtered through a 0.45 μm microporous membrane to obtain component B. Component B was loaded onto the antibody affinity chromatography column (HiTrap TM MabSelect TM ). After equilibrating the chromatography column with PBS, after the flow-through peak was balanced to the baseline, the affinity chromatography eluent (200 mM Gly-HCl, pH 2.7) was used for elution. Collect the elution peaks, add 1 / 10 volume of neutralizing solution (1M Tris-HCl pH9.0) to the elution peaks to neutralize the pH, and after ultrafiltration and concentration, use molecular sieves to remove endotoxins and determine the protein concentration. The collected hum...

Embodiment 2

[0060] This embodiment provides a method for preparing a mouse anti-human IgG monoclonal antibody by immunizing mice with the human IgG antigen obtained in Example 1, comprising the following steps:

[0061] Take 5 Balb / C mice and immunize them for the first time on the first day. Mix the human IgG antibody obtained in Example 1 with an equal amount of complete Freund's adjuvant. Rats were injected subcutaneously with a total injection volume of 200 μl.

[0062] The second immunization was carried out on the 14th day. The human IgG antibody obtained in Example 1 was mixed with an equal amount of incomplete Freund's adjuvant, fully emulsified, and 5 to 5 mice were subcutaneously injected according to the injection amount of 100 μg / mouse. Volume 200 μl.

[0063]One week later, measure the potency by ELISA method. If the titer does not meet the fusion requirements, immunize once every 14 days according to the second immunization method, and measure the titer one week later. Thr...

Embodiment 3

[0066] This embodiment provides the method for coupling the mouse anti-human IgG monoclonal antibody obtained in Example 2 to microspheres, comprising the following steps:

[0067] In this embodiment, the selected particle size is 6 μm and the density is 1.05 g / cm 3 840 μL of MES with a pH of 6.0, 80 μL of NHS with a concentration of 50 mg / mL, and 4 μL with a concentration of 50 mg / mL After mixing, let stand at room temperature for 15 minutes to obtain component C. This step can activate the carboxylic acid groups on the surface of the microspheres to attach NHS esters. Component C was centrifuged at 10,000 g at 4°C for 5 min, and the supernatant was discarded to obtain Component D. Component D was resuspended with 1 mL of PBS, and the water bath was sonicated for 3 minutes to obtain component E. Add 60 μL of the monoclonal anti-human IgG antibody obtained in Example 2 to Component E, mix well, shake gently at room temperature for 4 hours to obtain Component F, and this step...

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Abstract

The invention relates to the technical field of pharmaceutical preparations, in particular to a preparation method of an anti-platelet antibody detection signal preparation, the anti-platelet antibody detection signal preparation and application, the preparation method comprises the following steps: taking microspheres or particles which are uniformly coated with monoclonal anti-human IgG antibodies and can be distinguished in color by naked eyes and a composite stabilizer according to a formula ratio; and uniformly dissolving with a PBS solution to obtain the detection signal preparation. According to the preparation method of the anti-platelet antibody detection signal preparation, the microspheres of which the colors can be distinguished by naked eyes are used for replacing red blood cells, so that the preservation time can be greatly prolonged under the action of the compound stabilizer, synchronous purchase, storage and use with other components of a detected product are realized, and the stable preservation period can reach more than one year; and great convenience is provided for the production and transportation processes of anti-platelet antibody detection products.

Description

technical field [0001] The invention relates to the technical field of pharmaceutical preparations, in particular to a preparation method of an anti-platelet antibody detection signal preparation, an anti-platelet antibody detection signal preparation and its application. Background technique [0002] Platelet transfusion can be used to prevent and treat bleeding symptoms in patients with decreased platelet count or function deficiency, and to restore and maintain the normal hemostasis and coagulation function of the human body. However, long-term platelet transfusion may lead to serious consequences of ineffective platelet transfusion, that is, two consecutive transfusions of sufficient platelets did not improve the patient's clinical bleeding symptoms, and the platelet count did not increase significantly. Platelet antigens through blood transfusion, pregnancy and bone marrow transplantation can stimulate the body to produce anti-platelet antibodies to sensitize and destro...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/577G01N33/543G01N33/531
Inventor 严俊师丽佳刘涛蒋友红卿小红
Owner 宝锐生物科技泰州有限公司
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