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Preparation of polyclonal preparations for covid-19 patient-specific immunotherapy

A technology for COVID-19 and immunotherapy, applied in the field of preparation of polyclonal preparations, can solve the problems of not being able to inject finished products into the human body, containing toxic substances, expensive reagents, etc., to ensure high activity and quality, strict condition control, and reagents. Streamlined effect

Active Publication Date: 2021-05-11
广东龄值生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, the reagents used to prepare this polyclonal preparation are very expensive, difficult to popularize and use, and can only be used experimentally in laboratory theory, and there is no real relevant and practical product at present. More importantly, the preparation process is complex and rigorous. It is harsh, and various complex and even toxic substances are used, which makes the prepared polyclonal preparation contain more impurities and toxic substances, so it cannot be directly injected into patients for treatment
For example, in routine experiments, Phytohaemgglutin PHA is usually used to stimulate the emulsification of B lymphocytes, which can be used in the laboratory, but we cannot inject the finished product containing this substance into the human body, because PHA is toxic, which is also the world One of the substances whose use in the human body is banned by bodies such as the World Health Organization

Method used

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  • Preparation of polyclonal preparations for covid-19 patient-specific immunotherapy
  • Preparation of polyclonal preparations for covid-19 patient-specific immunotherapy
  • Preparation of polyclonal preparations for covid-19 patient-specific immunotherapy

Examples

Experimental program
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Effect test

Embodiment 1

[0035] Embodiment 1: the preparation method of the polyclonal preparation for COVID-19 patient-specific immunotherapy, comprises the following steps:

[0036] S1. Take 45ml of blood from recovered patients with COVID-19, add 3800g heparin, divide into 5 test tubes, use StemSep TM or EasySep TM CD19 mix to isolate CD19+ B lymphocytes from PBMNCs (peripheral blood mononuclear cells) by negative selection;

[0037] S2. Cultivate the CD19+B lymphocytes separated from step S1 with ACE solution. The culture period is 28 days. During this period, check the endotoxin in the culture medium every day, clean the culture medium and obtain the supernatant, and discard any indications Contaminated samples;

[0038] S3. When the supernatant collected in step S2 is freeze-dried and centrifuged with a centrifugal force of 1000g, the centrifugation time is 5 minutes, then the supernatant is obtained, concentrated by freeze-drying, and concentrated to the original volume by freeze-drying 1 / 4,...

Embodiment 2

[0041] Embodiment 2: the preparation method of the polyclonal preparation for COVID-19 patient-specific immunotherapy, comprises the following steps:

[0042] S1. Take 55ml of blood from a recovered patient with COVID-19, add 4200U heparin, divide it into 5 test tubes, and use StemSep TM or EasySep TM CD19 mix to isolate CD19+ B lymphocytes from PBMNCs by negative selection;

[0043] S2. Cultivate the CD19+B lymphocytes separated from step S1 with ACE solution. The culture period is 35 days. During this period, check the endotoxin in the culture medium every day, clean the culture medium and obtain the supernatant, and discard any indications Contaminated samples;

[0044] S3. When the supernatant collected in step S2 is freeze-dried and centrifuged with a centrifugal force of 1000g, the centrifugation time is 5 minutes, then the supernatant is obtained, concentrated by freeze-drying, and concentrated to the original volume by freeze-drying 1 / 4, and the endotoxin test is ca...

Embodiment 3

[0047] Embodiment 3: the preparation method of the polyclonal preparation for COVID-19 patient-specific immunotherapy, comprises the following steps:

[0048] S1. Take 50ml of blood from recovered patients with COVID-19, add 4000U heparin, divide into 5 test tubes, use StemSep TM or EasySep TM CD19 mix to isolate CD19+ B lymphocytes from PBMNCs by negative selection;

[0049] S2. Cultivate the CD19+B lymphocytes separated from step S1 with ACE solution. The culture period is 30 days. During this period, check the endotoxin in the culture medium every day, clean the culture medium and obtain the supernatant, discard any indication Contaminated samples;

[0050] S3. When the supernatant collected in step S2 is freeze-dried and centrifuged with a centrifugal force of 1000g, the centrifugation time is 5 minutes, then the supernatant is obtained, concentrated by freeze-drying, and concentrated to the original volume by freeze-drying 1 / 4, and the endotoxin test is carried out aft...

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Abstract

The invention discloses a method for preparing a polyclonal preparation for COVID‑19 patient-specific immunotherapy, which belongs to the field of biomedicine technology and includes the following steps: S1, taking the blood of a recovered patient from COVID‑19, adding heparin, and using StemSep TM or EasySep TM CD19 mixture is used to isolate CD19+B lymphocytes from PBMNCs by negative selection; S2, culture the isolated CD19+B lymphocytes with ACE solution, check the endotoxin in the culture medium every day, wash and obtain the supernatant, and discard Contaminated samples; S3. Freeze-dry and centrifuge the collected supernatant to obtain the supernatant, and then concentrate it for endotoxin testing; S4. Merge the supernatant, centrifuge and collect the supernatant; S5. Bacteria detection and toxin detection are carried out on the supernatant, and the qualified supernatant is selected to obtain the finished product. A polyclonal preparation for specific immunotherapy for COVID-19 patients can be prepared, and the prepared polyclonal preparation is safe, non-toxic, and low in cost.

Description

technical field [0001] The invention relates to the technical field of biomedicine, in particular to a method for preparing a polyclonal preparation for specific immunotherapy. Background technique [0002] COVID-19, the new type of coronavirus pneumonia, referred to as "new crown pneumonia", the health of people all over the world, especially all medical researchers and practitioners, is seriously threatened by the virus that causes the disease, SARS-CoV-2. The FDA has declared the safety and efficacy of treating patients with plasma from convalescent patients, and the efficacy has been demonstrated through trials and research conducted on 10 COVID-19 patients. However, the number of recovered patients is limited, and the amount of plasma is extremely scarce. At present, this treatment plan can only be used in critical situations, which seriously restricts the implementation and implementation of immunotherapy treatment with plasma from recovered patients. It is also diffi...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/10C12N5/0781C12P21/00A61K39/42A61P31/14
CPCA61K2039/505A61P31/14C07K16/10C07K2317/14C07K2317/21C07K2317/76C07K2317/94C12N5/0635C12N2500/32C12N2509/00C12P21/00
Inventor 阿拉·阿布德卡瑞姆穆罕默德弗德马颖
Owner 广东龄值生物科技有限公司