Hydrogel, and preparation method and three-dimensional cell culture method thereof

A technology of three-dimensional cells and culture methods, applied in the field of three-dimensional cell culture, can solve the problems of transformation, difficult in vitro research, and changing results

Inactive Publication Date: 2020-08-07
江苏安泰康健康科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

These methods do not accurately mimic the complexity of the extracellular microenvironment and may si...

Method used

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  • Hydrogel, and preparation method and three-dimensional cell culture method thereof
  • Hydrogel, and preparation method and three-dimensional cell culture method thereof
  • Hydrogel, and preparation method and three-dimensional cell culture method thereof

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preparation example Construction

[0047] One of the objects of the present invention is to provide a method for preparing a hydrogel for three-dimensional cell culture, the preparation method comprising:

[0048] Step S1, preparing porcine decellularized dermis and / or porcine decellularized bladder;

[0049] Step S2, freeze-drying the porcine decellularized dermis and / or the porcine decellularized bladder;

[0050] Step S3, grinding the freeze-dried pig decellularized dermis and / or freeze-dried the pig decellularized bladder into fine particles;

[0051] Step S4, adding the fine particles into the pepsin digestion solution, stirring for 48h-72h to form an extracellular matrix scaffold; and

[0052] Step S5 , adjusting the pH value of the pepsin digestion solution, so that the pepsin is inactivated, and the proteins in the extracellular matrix scaffold are respectively recombined to form the hydrogel.

Embodiment 1

[0053] Embodiment 1, prepare porcine dermal tissue and porcine bladder tissue decellularization reagent

[0054] 1) Raw material collection, full-thickness skin and bladder of adult pigs were obtained from local slaughterhouses.

[0055] 2) Decellularization reagents include, but are not limited to: triple distilled water, double distilled water, peracetic acid (English abbreviation: PAA) solution, 0.25% trypsin solution, 3% hydrogen peroxide (H 2 o 2 ) solution, 70% ethanol solution, Triton X-100 solution and 1X phosphate buffered saline solution (abbreviation: 1XPBS).

[0056] The peracetic acid (PAA) solution is an ethanol solution with a concentration of 0.1% peracetic acid, and the content of ethanol is 4%. It is mixed according to the components and their volumes shown in Table 1.

[0057] Table 1: Based on the weight of porcine dermal tissue and / or porcine bladder tissue and the corresponding volume of PAA solution

[0058]

[0059] Taking the data in the first ro...

Embodiment 2

[0067] Example 2. Preparation of Reagents and Equipment for Enzymatic Digestion and Hydrogel Formation of Porcine Acellular Dermis and Porcine Decellularized Bladder

[0068] Enzymatic digestion and hydrogel formation reagents, including but not limited to: porcine pepsin, 0.1N hydrochloric acid, 0.1N sodium hydroxide, 10XPBS and 1XPBS.

[0069] Concentration 1mg / mL pepsin preparation, 10mg porcine pepsin and 10mL 0.1N HCl were mixed in a 25mL Erlenmeyer flask.

[0070] Enzyme digestion and hydrogel formation equipment: lyophilizer, 40-mesh screen, non-humidified incubator.

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Abstract

The invention discloses a hydrogel, and a preparation method and a three-dimensional cell culture method thereof. The preparation method of the hydrogel comprises the following steps: S1, preparing pig acellular dermis and/or pig acellular bladder; S2, freeze-drying the porcine acellular dermis and/or the porcine acellular bladder; S3, grinding the freeze-dried porcine acellular dermis and/or freeze-drying the porcine acellular bladder into fine particles; S4, adding the fine particles into pepsin digestive juice, and stirring for 48-72 hours to form an extracellular matrix scaffold; and S5, adjusting the pH value and the salt concentration of the pepsin digestive juice to inactivate the pepsin, and recombining the protein in the extracellular matrix scaffold to form the hydrogel.

Description

technical field [0001] The invention relates to the technical field of biomaterials, and more specifically relates to a hydrogel prepared based on decellularization, a preparation method thereof, and a three-dimensional cell culture method using the hydrogel. Background technique [0002] It has long been believed that the extracellular matrix (ECM) not only provides structural support but also regulates cell growth, survival, maturation, differentiation and development of resident cells. Although many components of the ECM are preserved in different tissue types, each tissue is thought to have a unique composition. [0003] Recently, ECM scaffolds derived from the ECM of various tissues, including skin, adipose, pericardium, heart, skeletal muscle, and liver, have been observed to contribute to constructive regeneration when used as biomaterials in in vivo and in vitro experiments. To shape or form tissues in place. In some cases, these ECM scaffolds have tissue-specific ...

Claims

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Application Information

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IPC IPC(8): C12P21/06C08J3/075C08L89/00C12N5/00
CPCC08J3/075C08J2389/00C12N5/0062C12N2533/50C12N2533/90C12P21/06
Inventor 孙晓娇张文平张红霞刘星陆重益
Owner 江苏安泰康健康科技有限公司
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