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Rapid extraction method of human body fat stem cells and special human fat tissue digestant for same

A technology of adipose tissue and human fat, applied in the field of cell engineering, can solve the problems of complex methods, slow passage and proliferation, and small number of stem cells, and achieve the effect of high survival rate

Active Publication Date: 2014-03-19
JIANGSU RE STEM BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The disadvantage of this method of cultivating stem cells is that the method is complicated, the number of stem cells extracted is small, the purity is not high, and the subculture proliferation is slow.

Method used

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  • Rapid extraction method of human body fat stem cells and special human fat tissue digestant for same

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Embodiment 1: Preparation of a human fat digester A for digesting human adipose tissue according to the present invention

[0020] 1) Take 900ml of Dulbecco’s phosphate buffered saline (Dulbecco’s PBS, Sigma, D-8537, the same below), and then add 90000U of penicillin and streptomycin.

[0021] 2) Adjust the pH value: use 5% NaHCO 3 The pH of the solution was adjusted to 7.2.

[0022] 3) Add high-efficiency digestive agent: Add 150mg of collagenase type IV, 500mg of trypsin-EDTA complex, 100mg of α1 antitrypsin and 5000μL of dimethyl sulfoxide into Dulbecco’s phosphate buffer in sequence, and use shaking or ultrasonic to assist dissolution.

[0023] 4) Then add the above-mentioned Dulbecco's phosphate buffer solution to 1000mL, that is, the human fat digester A prepared by taking the lowest dose of each reagent component.

Embodiment 2

[0024] Embodiment 2: Preparation of a human fat digester B for digesting human adipose tissue according to the present invention

[0025] 1) Take 900ml of Dulbecco's phosphate buffered saline (Dulbecco's PBS, Sigma, D-8537), and then add 90000U of penicillin and streptomycin.

[0026] 2) Adjust the pH value: use 5% NaHCO 3 The pH of the solution was adjusted to 7.2.

[0027] 3) Add high-efficiency digestive agent: Add 2500mg of collagenase type IV, 5000mg of trypsin-EDTA complex, 1500mg of α1 antitrypsin and 100000μL of dimethyl sulfoxide to Dulbecco’s phosphate buffer in sequence, and use shaking or ultrasonic to assist dissolution.

[0028] 4) Then add the above-mentioned Dulbecco's phosphate buffer solution to 1000mL, that is, the human fat digester B prepared by taking the highest dose of each reagent component.

Embodiment 3

[0029] Example 3: Preparation of a human fat digester C for digesting human adipose tissue according to the present invention

[0030] 1) Take 900ml of Dulbecco's phosphate buffered saline (Dulbecco's PBS, Sigma, D-8537), and then add 90000U of penicillin and streptomycin.

[0031] 2) Adjust the pH value: use 5% NaHCO 3 The pH of the solution was adjusted to 7.2.

[0032] 3) Add high-efficiency digestive agent: Add 750 mg of collagenase type IV, 2500 mg of trypsin-EDTA complex, 500 mg of α1 antitrypsin and 30000 μL of dimethyl sulfoxide into Dulbecco’s phosphate buffer in sequence, and use shaking or ultrasonic to assist dissolution.

[0033] 4) Then add the above-mentioned Dulbecco's phosphate buffer solution to 1000mL, that is, the human fat digester C prepared by taking the optimal dose of each reagent component.

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Abstract

The application discloses a rapid extraction method of human body fat stem cells and a special human fat tissue digestant for the same. The method comprises human fat tissue treatment, liquiform fat centrifuging, and human body fat stem cell extraction. The special human fatty tissue digestant used in the method comprises a collagenase IV type, a trypsin-EDTA (ethylene diamine tetraacetic acid) complex, antitrypsin and dimethyl sulfoxide. The rapid extraction method can sufficiently separate fat mesenchymal stem cells, and simultaneously protects the cells from damage.

Description

technical field [0001] The invention relates to the field of cell engineering, in particular to a rapid extraction method of human adipose stem cells and a special human adipose tissue digester. Background technique [0002] Adipose-derived stem cells (ADSCs) are a type of adult stem cells widely used in the fields of tissue engineering and regenerative medicine, and have the same multipotential differentiation potential as bone marrow mesenchymal stem cells. ADSCs showed fibroblast-like growth, abundant cytoplasm and nucleoli, arranged in parallel or swirl-like arrangement. Cell cycle analysis showed that 69% of cells were in G0 / G1 phase, 24% in S phase, and 8% in G2 / M phase. The cells were subcultured for 2-3 days in the presence of fetal bovine serum to double the cell proliferation. After multiple passages (10-20 passages), the cell proliferation rate did not slow down significantly. Senescent cells appeared in the cell population after 6 passages, and senescent cells ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0775
Inventor 焦阳
Owner JIANGSU RE STEM BIOTECH