Optimized culture medium, kit and culture method for human placenta-derived mesenchymal stem cells

A technology for optimizing medium and stromal cells, applied in the field of optimizing medium for human placenta-derived mesenchymal stem cells, can solve the problems of different culture systems, inability to obtain MSCs, etc., achieve significant exponential growth, reduce large-scale cell preparation and Production cost, effect of large colony formation ability

Pending Publication Date: 2022-01-11
北京吉中科生物技术有限公司
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  • Abstract
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AI Technical Summary

Problems solved by technology

[0005] (1) The isolation and characterization of placental MSCs need to be standardized to ensure the stability of the preparation process for clinical cell preparations. At the same time (2) due to the differences in the operator's dissection structure of the placenta, different tissue location processing, and MSC isolation methods As well as the difference in the culture system, the target number of MSCs cannot be obtained from the placenta tissue
At present, there is no method to effectively obtain a large number of placenta-derived mesenchymal stem cells that can be used for clinical treatment

Method used

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  • Optimized culture medium, kit and culture method for human placenta-derived mesenchymal stem cells
  • Optimized culture medium, kit and culture method for human placenta-derived mesenchymal stem cells
  • Optimized culture medium, kit and culture method for human placenta-derived mesenchymal stem cells

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Experimental program
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Effect test

Embodiment 1

[0065] 1. Collection of placental tissue

[0066] (1) After the fetus is delivered, the placenta is completely separated from the mother, and the placenta is fully washed with normal saline, and then the surface is quickly rinsed with 75% medical alcohol.

[0067] (2) Quickly put the placenta into a tissue preservation bag, add tissue preservation solution (DMEM containing 4,500 mg / mL glucose and antibiotic solution (0.1% gentamicin, 0.2% streptomycin, and 0.12% penicillin)) to fully infiltrate , into the placenta collection box.

[0068] (3) Store the tissue preservation bag in a tissue transport box at 4°C and transport it to the laboratory. Samples were processed within 2 to 4 hours after tissue collection, and all operations were performed in a sterile laboratory environment.

[0069] (4) Place the sample in a medical stainless steel tray in a biological safety cabinet. Using a needle and syringe, rinse the tissue surface several times with 4°C pre-cooled D-PBS to remov...

Embodiment 2

[0110] (1) The placental mesenchymal stem cell culture method of the present invention has the stability of repeated application

[0111] The present invention sets up three isolation and expansion groups of placental tissue mesenchymal stem cells from different tissue donor sources, conducts microscopic examination of cell morphology and calculation of the total amount of preparation in the middle and later stages of the experiment, and verifies whether the cultivation method of the present invention can be realized Stable isolation and expansion of placenta-derived mesenchymal stem cells.

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Abstract

The invention relates to an optimized culture medium, a kit and a culture method for human placenta-derived mesenchymal stem cells, and belongs to the technical field of cell culture. The culture medium comprises DMEM-low glucose, L-glutamine, beta-mercaptoethanol, a KnockoutTM serum substitute, a recombinant human FGF2 protein, a recombinant human FGF4 protein, a recombinant human PDGFAB protein, a recombinant human EGF protein, a recombinant human VEGF165A protein, a recombinant human HGF protein, a recombinant human TGF-beta 1 protein, nicotinamide mononucleotide and non-essential amino acids. By the optimized culture medium, the in vitro amplification efficiency of the placenta-derived MSC can be maximally improved, the difference between batches of cells is reduced, the clinically relevant cell quantity in a shorter time range is reached, and good phenotype consistency and three-line differentiation potential in multiple generations can be maintained.

Description

technical field [0001] The invention relates to the technical field of cell culture, in particular to an optimized medium, a kit and a culture method for human placenta-derived mesenchymal stem cells. Background technique [0002] Mesenchymal stem cells (MSCs) are candidates for cell therapy, and in most cases, therapeutic responses have been shown to be cell dose dependent. Most applications target MSC-mediated tissue repair or immunomodulation, and in many applications allogeneic MSCs are economically advantageous compared to autologous MSCs, allowing the large-scale production of large cell preparations from a single tissue source for use in more patient. Human term placental tissue is rich in MSCs and is a physically large tissue that is usually discarded after fetal birth. Placenta is an ideal starting material for the large-scale production of multicellular doses of allogeneic MSCs. [0003] During the history of MSC cell therapy development, preclinical and clinica...

Claims

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Application Information

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IPC IPC(8): C12N5/0775
CPCC12N5/0668C12N2500/32C12N2500/44C12N2501/115C12N2501/119C12N2501/11C12N2501/135C12N2501/165C12N2501/12C12N2501/15C12N2500/40C12N2509/00C12N2509/10
Inventor 雷欣华金倞梁磊
Owner 北京吉中科生物技术有限公司
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