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

CN113913375APending Publication Date: 2022-01-11北京吉中科生物技术有限公司
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Patent Information

Authority / Receiving Office
CN · China
Current Assignee / Owner
Publication Date
2022-01-11

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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.
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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...

Examples

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.