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MSC Growth Medium 2 (C-28009) is an optimized medium formulation with reduced serum content to allow for more standardized culture conditions (considerably lower lot to lot variation). You can replace MSC Growth Medium by MSC Growth Medium 2. Coating of culture vessels is not necessary. We recommend to plate the cells (hMSCs from bone marrow; adipose tissue; or umbilical cord) at 4;000 cells/cm².
Our MSC Growth Medium XF (Ready-to-use) provides a xeno-free culture system for human MSCs. It contains all growth factors and supplements except attachment- and spreading factors. Therefore; culture vessels must be precoated with 10 μg/ml human or bovine fibronectin.
Protocol: Fibronectin coating
Dilute the Fibronection Solution to 10 μg/ml final concentration in Dulbecco´s PBS w/o Calcium and Magnesium. Overlay the culture surface of your tissue culture vessel with an amount of the diluted Fibronectin Solution sufficient to effectively coat the complete surface. Be sure that the entire surface is covered. Place flasks on a level surface at RT for 60 min. Aspirate the excess Fibronectin Solution and use immediately or let air-dry the open vessel under a laminar flow bench. Unused vessels may be stored at 4°C for up to 2 weeks.
Usually 90-100% of the hMSC show a neuronal morphology after differentiation with our Mesenchymal Stem Cell Neurogenic Differentiation Medium (C-28015). 60-80% of them are positive for nissl bodies after a nissl stain. However the differentiation capacity depends on the origin of the cells and the number of population doublings they have undergone.
Differentiation of hMSC into mature adipocytes takes approx. 2 weeks. You can keep the adipocytes for up to 3 weeks in the MSC Adipogenic Differentiation Medium. After 3 weeks we recommend to switch to PromoCell's Adipocyte Nutrition Medium (C-27438).
Human Mesenchymal Stem Cells have retained the ability to differentiate into a variety of cell types including fat cells; chondrocytes; and osteoblasts. PromoCell supply hMSC from 3 different tissues: bone marrow (hMSC-BM; C-12974); umbilical cord matrix (hMSC-UC; C-12971); and adipose tissue (hMSC-AT; C-12977).
- MSC-BM show very good differentiation into bone cells but also into chondrocytes and fat cells when the respective Differentiation Media is used.
- MSC-UC have a high potential to differentiate into chondrocytes but only weak potential for fat or bone cell differentiation.
- In contrast; MSC-AT differentiate very well into fat and bone cells but only moderately into chondrocytes.
Our hMSC-AT are characterized by their differentiation potential into chondrocytes; fat cells; and bone cells. In addition; we determine the presence of CD73; CD90 and CD105 expression as well as the absence of CD14; CD19; CD34; CD45 & HLA-DR expression by flow cytometry as proposed by the International Society for Cellular Therapy.
Yes; we have received a customer feedback that our MSC Growth Medium 2 also works for rat MSCs.
The rat cells grow nicely in this medium and have a good viability.
Yes; you can use 4.5% neutral buffered formalin. Paraformaldehyde should work as well.
You can also use Oil Red O to stain lipid droplets. At PromoCell; we used to use Oil Red O as well; but switched to Sudan III some time ago for organizational reasons.
Yes; there are a few differences:
- We recommend replacing the MSC Growth Medium XF (C-28019) 3-4 h after seeding; as opposed to 16-24 hours after seeding for most other cell types/growth media.
- When MSC Growth Medium XF; MSC Neurogenic Differentiation Medium; MSC Adipogenic Differentiation Medium 2 or MSC Osteogenic Differentiation Medium are used; flasks have to be coated with 10 µg/cm² (human or bovine) fibronectin according to the instruction manual.
- We strongly recommend using Accutase (C-41310) for cell detachment instead of Trypsin. If Trypsin is used; contact time should not exceed 2 min.
The PBS buffering enhances the Alizarin Red staining (precipitation of the dye) and makes it more intense.
Leave the PBS on the cells after staining/washing and analyze the sample immediately; as the dye may bleed upon prolonged storage without embedding.
You could probably use our MSC Chondrogenic Differentiation Medium without inducers (C-28014). It is also serum-free and the same as C-28012 just without chondrogenic inducers.