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Primary separation culture method of umbilical vein endothelial cells and reagent box of primary separation culture method

A technology for the separation and culture of umbilical cord veins, which is applied to animal cells, vertebrate cells, artificial cell constructs, etc., and can solve the problems of inability to provide nutrients to enzymatically hydrolyzed cells, long primary culture period, and small number of endothelial cells. Achieve the effects of shortening the primary culture cycle, ensuring nutrient supply, and high cell viability

Inactive Publication Date: 2015-08-12
GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0013] (1) The single-enzyme enzymatic hydrolysis method is adopted, which takes a long time for enzymatic hydrolysis, and the number of endothelial cells obtained is small and their vitality is low;
[0014] (2) The culture system is only M199+10% FBS, with the addition of an appropriate amount of antibiotics, it cannot provide sufficient nutrients for the cells that have just been enzymatically hydrolyzed, and the primary culture period is too long;
[0015] (3) The enzymatic hydrolysis takes a long time, and different types of cells are obtained, which is not conducive to the purification of cells after passage

Method used

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  • Primary separation culture method of umbilical vein endothelial cells and reagent box of primary separation culture method
  • Primary separation culture method of umbilical vein endothelial cells and reagent box of primary separation culture method
  • Primary separation culture method of umbilical vein endothelial cells and reagent box of primary separation culture method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0070] Example 1 Primary Separation and Culture of Umbilical Cord Vein Endothelial Cells

[0071] Obtain the umbilical cords of full-term infants in the hospital, wash the umbilical cords with PBS containing 1× antibiotics, soak them in umbilical cord preservation solution, store them at 4°C and transport them, and enter the experimental procedure within 24 hours after delivery. Umbilical cord preservation solution: normal saline contains 1×~2× penicillin-streptomycin (double antibody), 50 mg / mL lacturonic acid, 5% hydroxyethyl starch, and 5% umbilical cord plasma.

[0072] Take out the vein and place it in a 15cm glass dish, wash it twice with PBS containing 1×antibiotics, cut off 1cm at both ends, transfer it to a new 15cm glass dish, and wash it once more with PBS containing 1×antibiotics. Use a 50mL disposable syringe to draw the lavage solution (lavage solution: PBS containing 1×~3× double antibody, 0.02% EDTA) and infuse it into the umbilical cord vein until the blood in...

Embodiment 2

[0084] Example 2 Primary Separation and Culture of Umbilical Cord Vein Endothelial Cells

[0085] Obtain the umbilical cords of full-term infants in the hospital, wash the umbilical cords with PBS containing 1× antibiotics, soak them in umbilical cord preservation solution, store them at 4°C and transport them, and enter the experimental procedure within 24 hours after delivery. Umbilical cord preservation solution: normal saline contains 1×~2× penicillin-streptomycin (double antibody), 100 mg / mL lactobionic acid, 1% hydroxyethyl starch, and 5% umbilical cord plasma.

[0086] Take out the vein and place it in a 15cm glass dish, wash it twice with PBS containing 1×antibiotics, cut off 1cm at both ends, transfer it to a new 15cm glass dish, and wash it once more with PBS containing 1×antibiotics. Use a 50mL disposable syringe to draw the lavage solution (lavage solution: PBS containing 1×~3× double antibody, 0.04% EDTA) and infuse it into the umbilical cord vein until the blood ...

Embodiment 3

[0091] Example 3 Primary Separation and Culture of Umbilical Cord Vein Endothelial Cells

[0092] Obtain the umbilical cords of full-term infants in the hospital, wash the umbilical cords with PBS containing 1× antibiotics, soak them in umbilical cord preservation solution, store them at 4°C and transport them, and enter the experimental procedure within 24 hours after delivery. Umbilical cord preservation solution: normal saline contains 1×~3×penicillin-streptomycin (double antibody), 80mg / mL lacturonic acid, 2% hydroxyethyl starch, and 5% umbilical cord plasma.

[0093] Take out the vein and place it in a 15cm glass dish, wash it twice with PBS containing 1×antibiotics, cut off 1cm at both ends, transfer it to a new 15cm glass dish, and wash it once more with PBS containing 1×antibiotics. Use a 50mL disposable syringe to draw the lavage solution (lavage solution: PBS containing 1×~3× double antibody, 0.04% EDTA) and infuse it into the umbilical cord vein until the blood in t...

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Abstract

The invention relates to the technical field of stem cell separation culture, in particular to a primary separation culture method of umbilical vein endothelial cells and a reagent box of the primary separation culture method. The method includes: filling enzymatic hydrolysate into an umbilical vein for enzymolysis, and collecting the umbilical vein endothelial cells; using a complete culture medium to culture the umbilical vein endothelial cells, wherein the enzymatic hydrolysate is composed of a basic culture medium containing II-type collagnase and hyaluronidase. The primary separation culture method is simple in operation step, high in cell acquiring rate and high in cell survival rate, so that primary culture period is shortened greatly.

Description

technical field [0001] The invention relates to the technical field of stem cell separation and culture, in particular to a primary separation and culture method of umbilical cord vein endothelial cells and a kit thereof. Background technique [0002] A thrombus is a small piece of blood that forms on the surface of a vessel in the cardiovascular system where the lining has been peeled off or repaired. In the variable fluid-dependent form, the thrombus consists of insoluble fibrin, deposited platelets, accumulated leukocytes, and trapped erythrocytes. Thrombosis is a multifactorial change process in which a group of genetic and environmental factors interact and influence each other. The main characteristics of common clinical thrombosis patients are familial heredity, recurrence, severity of symptoms, abnormal thrombosis site, and younger onset time. Formation mechanisms include: [0003] 1. Intimal injury of heart and blood vessels [0004] (1) When the intima is damag...

Claims

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Application Information

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IPC IPC(8): C12N5/071
Inventor 葛啸虎陈海佳王一飞卢瑞珊王小燕李平
Owner GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD