Fecal microbiota collection, cryopreservation and unfreezing method of fecal microbiota transplantation technology

A technology of fecal bacteria transplantation and fecal bacteria, which is applied in the field of freezing and thawing, and the collection of fecal bacteria. It can solve the problems of long storage and freezing operation time, low activity rate, and high technical cost of fecal bacteria, so as to shorten the thawing time and improve the survival rate. , high freezing efficiency

Active Publication Date: 2020-11-03
山东省大健康精准医疗产业技术研究院
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] Aiming at the problems of long storage and freezing operation time of fecal bacteria, low activity rate after thawing and high technical cost in the prior art

Method used

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  • Fecal microbiota collection, cryopreservation and unfreezing method of fecal microbiota transplantation technology
  • Fecal microbiota collection, cryopreservation and unfreezing method of fecal microbiota transplantation technology
  • Fecal microbiota collection, cryopreservation and unfreezing method of fecal microbiota transplantation technology

Examples

Experimental program
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Effect test

Embodiment 1

[0047] A method for collecting, freezing and thawing fecal bacteria of a fecal bacteria transplantation technique, comprising the following steps:

[0048] (1) Preparation of fecal bacteria: put the fresh feces of the donor in a nitrogen bioengineering kitchen, weigh about 50 g and put it in a blender, add 250 mL of sterile 0.85% NaCl solution for the first homogenization treatment, and mix the feces slurry The body is filtered through a 5mm stainless steel filter to remove the large particles; then add 250mL sterile 0.85% NaCl solution in the homogenizer for the second homogeneous treatment to obtain the fecal bacteria slurry; then pass the fecal bacteria slurry step by step Filter through 2.0mm, 1.0mm, 0.5mm and 0.25mm stainless steel filters to remove unabsorbed food residues and small particulate matter; centrifuge at 4000 rpm for 10 minutes to obtain 5g of fecal bacteria;

[0049] (2) Vitrification: At 25°C, add 9 mL of pretreatment liquid to the fecal bacteria obtained i...

Embodiment 2

[0061] A fecal bacteria collection, freezing and thawing method for fecal bacteria transplantation technology, comprising the following steps:

[0062] (1) Preparation of fecal bacteria: put the fresh feces of the donor in a nitrogen bioengineering kitchen, weigh about 50g and place it in a blender, add 250mL of sterile 0.85% NaCl solution for the first homogenization treatment, and put the feces The slurry was filtered through a 7mm stainless steel filter to remove large particles; then 250mL of sterile 0.85% NaCl solution was added to a homogenizer for the second homogeneous treatment to obtain the fecal bacteria slurry; then the fecal bacteria slurry was graded Filter through 2.0mm, 1.0mm, 0.5mm and 0.25mm stainless steel filter screens to remove unabsorbed food residues and small particulate matter; centrifuge at 5000 rpm for 12min to obtain 8g of fecal bacteria;

[0063] (2) Vitrification: Add 14 mL of pretreatment solution to the fecal bacteria obtained in step (1) at 25...

Embodiment 3

[0075] A fecal bacteria collection, freezing and thawing method for fecal bacteria transplantation technology, comprising the following steps:

[0076] (1) Preparation of fecal bacteria: put the fresh feces of the donor in a nitrogen bioengineering kitchen, weigh about 50g and place it in a blender, add 250mL of sterile 0.85% NaCl solution for the first homogenization treatment, and put the feces The slurry was filtered through a 10mm stainless steel filter to remove large particles; then 250mL of sterile 0.85% NaCl solution was added to a homogenizer for a second homogeneous treatment to obtain the fecal bacteria slurry; then the fecal bacteria slurry was graded Filter through 2.0mm, 1.0mm, 0.5mm and 0.25mm stainless steel filter screens to remove unabsorbed food residues and small particulate matter; centrifuge at 6000 rpm for 15 minutes to obtain 11g of fecal bacteria;

[0077] (2) Vitrification: at room temperature, add 20 mL of pretreatment liquid to the fecal bacteria ob...

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Abstract

The invention relates to a fecal microbiota collection, cryopreservation and unfreezing method of a fecal microbiota transplantation technology. The method comprises the following steps: (1) preparingfecal microbiota; (2) carrying out vitrification freezing on the fecal microbiota prepared in the step (1) to obtain cryopreserved fecal microbiota; and (3) unfreezing and recovering the cryopreserved fecal microbiota obtained in the step (2). According to the invention, a vitrification freezing technology is adopted, so that the freezing rate (greater than 20000 DEG C / min) is greatly increased,cells rapidly form a 'glassy state' in a rapid cooling process, ice crystals are prevented from being formed inside and outside the cells, excessive damage to florae in the fecal microbiota is avoided, and an antifreezing protein is added into a cryoprotectant, so that the cytotoxicity of the cryoprotectant is avoided; and the survival rate of the flora in the unfrozen fecal microbiota liquid is increased to 90% or above.

Description

technical field [0001] The invention relates to a method for collecting, freezing and thawing feces bacteria in the technology of fecal bacteria transplantation, and belongs to the technical field of fecal bacteria transplantation. Background technique [0002] Fecal microbiota transplantation (FMT) is a microecological treatment method that isolates functional flora from the feces of healthy donors and transplants them into the digestive tract of patients to rebuild the intestinal flora of patients. It is used in the treatment of various gastrointestinal and metabolic diseases, such as chronic functional constipation, refractory Clostridium difficile infection, inflammatory bowel disease, etc. [0003] Tracing back to the origin of FMT, the earliest written record of its use in treating human diseases is "Elbow Reserve Emergency Prescription" (also known as "Elbow Back") written by Ge Hong in the Eastern Jin Dynasty (300-400 AD), which describes the use of human feces to tr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/04C12N1/02C12N1/20C12R1/01
CPCC12N1/04C12N1/02C12N1/20
Inventor 高丽鹤汪运山杨炜华朱之炜李文靖苏真真张绍明
Owner 山东省大健康精准医疗产业技术研究院
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