Why is PBS added in the last step of the Alizarin Red staining protocol (Osteogenic differentiation and analysis of MSC)? Should the PBS be aspirated before analysis?

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.

I have accidentally stored the Cryo-SFM at -20°C. Can the product still be used in this case?

According to the product manual, Cryo-SFM should be stored at 4-8°C. However, since this solution is used to freeze cells in liquid nitrogen, we assume that storing Cryo-SFM once at -20°C should not have a negative impact on the product quality. After thawing, please store it at 4-8°C as recommended.

How can I avoid precipitates when preparing my Mesenchymal Stem Cell Adipogenic Differentiation Medium 2?

The supplement should be at room temperature when added to the MSC Adipogenic Basal Medium 2. It may also be beneficial to invert the tube a few times to bring precipitates back into solution. Please note: It is not recommended to filter the basal medium, supplements, or complete medium, as components that induce or promote differentiation may be removed, resulting in a low differentiation rate when using the medium.

Are there any differences in the cultivation protocol when cultivating MSCs compared to other cell types?

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, including MSC Growth Medium 2. IF MSC Growth Medium XF, MSC Neurogenic Differentiation Medium, MSC Adipogenic Differentiation Medium 2, or MSC Osteogenic Differentiation Medium are used, the TC vessels must be pre-coated according to the instruction manual. We strongly recommend using Accutase (C-41310) for cell detachment instead of Trypsin. If Trypsin is used, the contact time should not exceed 2 min.

After thawing the supplements, I see some precipitation. Is this normal?

Light flocculation may be seen upon thawing the supplements containing ECGS/heparin or BPE. This does not affect the activity of our media. Optionally, the precipitate can be removed by centrifugation under sterile conditions. We recommend to thaw the supplements (SupplementMix or SupplementPack) at 15-25°C.

Are PromoCell HPAEC and HPASMC obtained from the proximal or distal part of the pulmonary artery?

Our HPAEC and HPASMC are isolated from the proximal pulmonary artery.

Why are PromoCell products "for in vitro research use only"?

The basis for the intended use of our products is defined in our Terms & Conditions under the chapter “Use of Goods”.

How long do I need to activate my macrophages? Can I stop adding activation factors after 24 hours?

The activation of macrophages as such is complete after 24 hrs. However, to maintain the activation status over a longer period of time (i.e., several days), fresh activation factors should be added with every medium change.

Can the Cytokine Mix E for the Hematopoietic Progenitor Expansion Medium XF (C-39891; 5 ml) be safely aliquoted?

Yes, it is possible to aliquot the 5 ml Cytokine Mix E (C-39891) into 5 x 1 ml.

What is the difference between PromoCell NHEK and NHEK GM3?

The NHEK (primary human keratinocytes) are isolated in our serum-free Keratinocyte Growth Medium 2 (C-20011), the NHEK GM3 in our improved serum-free and BPE-free Keratinocyte Growth Medium 3 (C-20021). Both, NHEK and NHEK GM3 are available from single or from pooled donors isolated from the epidermis of juvenile foreskin or adult skin.

Will my NHEK (isolated in PromoCell Keratinocyte Growth Medium 2) grow in PromoCell Keratinocyte Growth Medium 3?

Yes, the NHEK-GM2 (C-12001, C-12003, C-12005, C-12006) also grow in PromoCell Keratinocyte Growth Medium 3 (C-20021). Using the protocol with the fixed intervals, they grow slightly faster than in Keratinocyte GM2 and proliferate for > 15 PDs. Conversely, NHEK-GM3 (C-12011, C-12013, C-12015, C-12016) also grow in the existing Keratinocyte Growth Medium 2 (C-20011). When using the classical subcultivation protocol (density > 70% - 90%), they grow slightly slower compared to Keratinocyte GM3, but also reach > 15 PDs.

Technical library - RESOURCES

The use of antibiotics creates a false sense of security and allows users to develop poor aseptic techniques. This leads to low-level contamination with partially resistant bacteria occurring but being overlooked for a time. This then leads to cells with undetected contamination being cultured for extended periods of time, increasing the risk that contamination will spread throughout the laboratory and eventually antibiotic-resistant strains of bacteria may develop. Mycoplasma infections can also occur more easily, as they are often introduced along with contaminants such as bacteria and fungi. Last but not least, antibiotics are known to have negative effects on the metabolism of eukaryotic cells - more details on this topic can be found in our blog article "Antibiotics in cell culture: friend or enemy".

Further Information

Blog article: Antibiotics in Cell Culture: Friend or Enemy?