Compound enzyme digestive juice as well as preparation method and application thereof

A digestive juice and compound enzyme technology, applied in the field of compound enzyme digestive liquid and its preparation, can solve the problems of reduced cell activity, morphological rupture, single composition, etc., and achieve the effects of low cell death rate, short digestion time and convenient operation

Active Publication Date: 2021-03-12
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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  • Compound enzyme digestive juice as well as preparation method and application thereof
  • Compound enzyme digestive juice as well as preparation method and application thereof
  • Compound enzyme digestive juice as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
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 digestive juice as well as a preparation method and an application thereof. The digestive juice comprises the following components with final concentrations: 100-200U / mL of type IV collagenase, 1-10U / ml of type II dispersive enzyme, 50-200U / ml of deoxyribonuclease I, 0.1-0.5 mM of PMSF, 10wt%-20wt% of ethylene glycol, 5wt%-10wt% of polysucrose, and sterilewater serving as a solvent, wherein the pH value of the digestive juice is 7.4-8.0. The preparation method comprises the following steps: preparing materials according to the final concentration of each component of the digestive juice; dissolving type IV collagenase, type II dispersive enzyme, deoxyribonuclease I, PMSF and polysucrose in sterile water, adding ethylene glycol, and uniformly mixing; and adjusting the pH value to 7.4-8.0, filtering and sterilizing to obtain the product. The compound enzyme digestive juice and the preparation method thereof are applied to the organoids technology. The digestive juice disclosed by the invention not only can be used for obtaining complete organoid, but also can be used for obtaining cell clusters with uniform dispersion sizes, and is short in digestion time, convenient to operate and stable in 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 Applications(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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