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A kind of compound enzyme digestive liquid and its preparation method and application

A technology of digestive juice and compound enzyme, which is applied in the field of compound enzyme digestive liquid and its preparation, can solve the problems of reduced cell activity, impact on experiments, and morphological rupture, and achieve the effects of low cell death rate, convenient operation, and short digestion time

Active Publication Date: 2022-04-12
ACCURATE INT BIOTECHNOLOGY (GUANGZHOU) CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The conventional separation method is to use trypsin to digest. However, because the volume of organoids is micron-scale and the composition is single, they are relatively fragile. During the digestion process of the existing trypsin digestion system, they are prone to morphological rupture and reduced cell activity. situation, thus affecting the subsequent experiments

Method used

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  • A kind of compound enzyme digestive liquid and its preparation method and application
  • A kind of compound enzyme digestive liquid and its preparation method and application
  • A kind of compound enzyme digestive liquid and its preparation method and application

Examples

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Effect test

Embodiment 1

[0052] This embodiment provides a compound enzyme digestion solution, including the following components at the final concentration: type IV collagenase 200U / mL, type II dispase 2U / ml, deoxyribonuclease 20U / ml, PMSF 0.1mM, ethylene glycol 10 wt%, polysucrose 10 wt%, the solvent is sterile water; the pH of the digestive juice is 7.4. The preparation method of the digestive juice comprises: batching ingredients according to the final concentration of each component of the digestive juice; Mix the diol evenly; adjust the pH to 7.4, filter and sterilize, and store the digestive juice in a -20°C refrigerator for later use.

Embodiment 2

[0054] This embodiment provides a compound enzyme digestion solution, which is different from Embodiment 1 in that it also includes: CaCl 2 2mM, MgCl 2 1mM. The preparation method of the digestive juice comprises: batching ingredients according to the final concentration of each component of the digestive juice; 2 , MgCl 2 1. Dissolve polysucrose in sterile water, then add ethylene glycol and mix evenly; adjust the pH to 7.4, filter and sterilize, and store the digestive juice in a -20°C refrigerator for later use.

Embodiment 3

[0056] This embodiment provides a compound enzyme digestion solution, which differs from that of Embodiment 1 in that it also includes: 1 wt% glycerin. The preparation method of the digestive juice comprises: batching ingredients according to the final concentration of each component of the digestive juice; Glycol and glycerin are mixed evenly; adjust the pH to 7.4, filter and sterilize, and store the digestive juice in a -20°C refrigerator for later use.

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Abstract

The invention provides a compound enzyme digestion liquid and its preparation method and application. The digestive solution includes the following final concentrations of components: type IV collagenase 100‑200U / mL, type II dispase 1‑10U / ml, deoxyribonuclease Ⅰ 50‑200U / ml, PMSF 0.1‑0.5mM, ethylene glycol Alcohol 10wt%-20wt%, polysucrose 5wt%-10wt%, solvent is sterile water; pH of the digestive juice is 7.4-8.0. The preparation method includes: mixing ingredients according to the final concentration of each component of the digestive juice; first dissolving type IV collagenase, type II dispase, deoxyribonuclease I, PMSF and polysucrose in sterile water, and then adding ethylene glycol to mix Uniform; adjust the pH to 7.4‑8.0, and filter to sterilize. The composite enzyme digestion liquid and its preparation method are applied in organoid technology. The digestive solution of the invention can obtain both complete organoids and dispersed cell clusters of uniform size, and has short digestion time, convenient operation and stable effect.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to a compound enzyme digestive liquid and its preparation method and application. Background technique [0002] Organoids are organ-specific collections of cells derived from stem cells or precursor cells. Different from a single cell group that is routinely cultured in a two-dimensional environment, organoid culture is a variety of cell groups contained in a specific organ in a three-dimensional environment. It can even be said that organoids are the best in vitro drug screening models. [0003] Because organoid culture is based on Matrigel droplets, if organoids need to be collected for passage cryopreservation or other downstream applications, the first thing to do is to completely separate the organoids from the gel. The conventional separation method is to use trypsin to digest. However, because the volume of organoids is micron-scale and the composition is si...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/00C12N5/071
CPCC12N5/00C12N5/0602C12N2509/00
Inventor 谢素瑶朱宇廖传荣黄敏
Owner ACCURATE INT BIOTECHNOLOGY (GUANGZHOU) CO LTD
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