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A kind of extracorporeal liver perfusion device and its application

A technology of perfusion and perfusate, applied in biochemical cleaning devices, enzymology/microbiology devices, bioreactors/fermenters for specific purposes, etc. The cost of hepatocytes, difficulty in controlling the temperature of the perfusate, etc., can save manpower, improve vitality, and maintain a constant temperature.

Active Publication Date: 2017-01-25
WEST CHINA HOSPITAL SICHUAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For large animals, such as pigs, dogs, and human livers, due to the large volume, large amount of perfusate, and long time, no matter in situ or in vitro perfusion, traditional artificial syringe injection perfusion cannot be used, and perfusion pumps can only be used.
However, when using a perfusion pump for perfusion, due to the lack of a dedicated complete set of equipment, it is very easy to cause contamination such as bacteria during the perfusion separation process.
Collagenase has high requirements on temperature, 37°C is its optimum temperature, and it is difficult to control the temperature of the perfusate without special perfusion equipment. Therefore, the final cell digestion effect is often affected due to poor temperature control, which cannot be achieved. Required cell quantity and quality requirements, and cause waste of expensive collagenase, increase the cost of isolating and culturing hepatocytes

Method used

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  • A kind of extracorporeal liver perfusion device and its application
  • A kind of extracorporeal liver perfusion device and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] Example 1 The extracorporeal liver perfusion device of the present invention

[0060] Such as figure 1 The extracorporeal liver perfusion device shown includes a perfusate I storage container 1 with a volume of 10 L. The upper end of the storage container 1 has a liquid inlet funnel 11, and a liquid sterilizing filter membrane 14 is placed at the bottom of the funnel 11. The upper end of the storage container 1 has a An air vent 17 is connected with an air filter 18, a liquid level gauge 23 is arranged on the side wall of the storage container 1, a liquid outlet 26 is arranged at the lower end of the storage container 1, and a stop valve 29 is arranged at the outlet. The device also includes a perfusate II storage container 2 with a volume of 2L. There is a liquid inlet funnel 12 at the upper end of the storage container 2. A liquid sterilizing filter membrane 15 is placed at the bottom of the funnel 12. There is a vent hole 19 at the upper end of the storage container ...

Embodiment 2

[0072] Example 2 The method for the separation and preparation of primary hepatocytes using the extracorporeal liver perfusion device of the present invention

[0073] 1. Test reagents

[0074] Perfusate I, perfusate II, perfusate III, liver cell washing solution;

[0075] The respective preparation methods are as follows:

[0076] 1. Perfusate Ⅰ: Take the preparation of 1 liter reagent as an example:

[0077] (1) Take the following ingredients: 8.3 grams of sodium chloride, 0.5 grams of potassium chloride, 2.4 grams of HEPES, 4 grams of NAC, and 0.95 grams of EGTA;

[0078] (2) Mix the above ingredients, dissolve in deionized water, stir at room temperature for 10 minutes to fully dissolve, adjust the volume to 1L, adjust the pH to 7.4, filter and sterilize with a 0.2 micron filter membrane, and store at room temperature. When in use, preheat to 37°C.

[0079] 2. Perfusate II: Take the preparation of 1 liter reagent as an example:

[0080] (1) Take the following ingredie...

Embodiment 3

[0100] Example 3 The method of using the extracorporeal liver perfusion device of the present invention to separate and prepare primary hepatocytes

[0101] 1. Test reagents

[0102] 1. Perfusate Ⅰ: Take the preparation of 1 liter reagent as an example:

[0103] (1) Take the following ingredients: 8.1 grams of sodium chloride, 0.5 grams of potassium chloride, 2.6 grams of HEPES, 4.5 grams of NAC, and 1.1 grams of EGTA;

[0104] (2) Mix the above ingredients, dissolve in deionized water, stir at room temperature for 10 minutes to fully dissolve, adjust the volume to 1L, adjust the pH to 7.4, filter and sterilize with a 0.2 micron filter membrane, and store at room temperature. When in use, preheat to 37°C.

[0105] 2. Perfusate II: Take the preparation of 1 liter reagent as an example:

[0106] (1) Take the following ingredients: 8.1 grams of sodium chloride, 0.5 grams of potassium chloride, and 2.6 grams of HEPES;

[0107] (2) Mix the above ingredients, dissolve in deioniz...

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Abstract

The invention discloses an in-vitro liver perfusion device which comprises a perfusion liquid storage container set composed of 3 perfusion liquid storage containers, a four-way pipe, a perfusion pump, a heat exchange system, an oxygenator, a defoaming device, a perfusion catheter, a three-way pipe and a reflux pump. The in-vitro liver perfusion device can perform three-step perfusion on the in-vitro liver, maximally perform effective perfusion on the pre-perfusion liquid and digestive juice quickly and continuously, and maximally control the hepatic tissue digestion process, thereby saving the cost, keeping the temperature constant, reducing the pollution, and further obtaining sufficient experimental and clinical liver cells with high quality (especially abundant liver cells required by liver cell transplantation, biological artificial livers and the like).

Description

technical field [0001] The present invention relates to an extracorporeal liver perfusion device and its application. Background technique [0002] Primary porcine hepatocytes are the most suitable source of heterogeneous hepatocytes in the treatment of BAL. The method used to separate mature hepatocytes is enzymatic digestion. This method is to inject collagenase into the blood vessels of the liver and act on the liver tissue to make The complex fibrous junctions between hepatocytes are dissolved, thereby obtaining intact individual hepatocytes. For large animals, such as pigs, dogs, and human livers, due to the large volume, large amount of perfusate, and long time, no matter in situ or in vitro perfusion, traditional manual syringe injection perfusion cannot be used, and a perfusion pump can only be used. However, when the perfusion pump is used for perfusion, due to the lack of special complete sets of equipment, it is very easy to cause bacterial and other pollution du...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12M1/34C12M1/02C12N5/071
CPCC12M23/58C12M33/14C12M41/18C12N5/067
Inventor 包骥步宏石毓君
Owner WEST CHINA HOSPITAL SICHUAN UNIV
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