Technical library

Please find attached a trouble shooting guide to identify possible reasons for poor detachment during subculture.

Microbiological contaminations can be introduced into cell cultures by unsterile working techniques, contaminated water baths, media, plasticware, etc. Please check the attached trouble shooting guide for more detailed information.

Your observation can be either ascribed to a change of the culture medium, to a cross-contamination with another cell type, or to differentiation or senescence. The attached trouble shooting guide should help you to identify the reason for your observation.

Normal cells have a finite life span and therefore eventually stop growing and become senescent. In addition, a cease in proliferation or cell death can be induced by other factors. For more details, check the trouble shooting guide below.

In principle, all microvascular endothelial cells should be able to migrate, proliferate and form tubes or sprouts in an appropriate assay after angiogenic stimulation. As this is not part of our routine quality control procedure, we cannot tell for sure whether all cell lots will respond to angiogenic stimuli. However PromoCell also supplies HDMEC pre-screened (C-12215) that are especially tested for a positive VEGF response.

No, we don't determine the ratio of HDLEC and HDBEC in our HDMEC lots. From our experience, the percentage of HDLEC is highly lot-dependent and can vary between 5 and 60%.

Our HDMEC are isolated from the dermis of juvenile foreskin or adult skin. The purity is > 95%. Since the dermis contains blood and lymphatic capillaries, HDMEC cultures comprise blood and lymphatic microvascular endothelial cells that have differing morphologies. Both cell types have a common origin and can be identified by several markers. The ratio of lymphatic and blood derived endothelial cells can vary from lot to lot and is not determined at PromoCell.

• FBS (or BSA) can be substituted for HSA, however we do not recommend this as the FBS will lead to unpreferable immunologic stimulation of human macrophages.

• The addition of EDTA to the PBS is to further augment the “anti-clumping” activity of the Ca²⁺/Mg²⁺ free PBS.

A concentration > 10% FBS is needed to completely inactivate the trypsin. As most of PromoCell's growth media are serum-reduced or serum-free, the use of a trypsin inhibitor like TNS is highly recommended.

The standard medium for isolation and propagation of our HUVEC, HUAEC, HPAEC, and HSaVEC is Endothelial Cell Growth Medium (C-22010). It contains ECGS, an extract from bovine hypothalamus which has mitogenic effects on endothelial cell proliferation. Scientists who prefer a more defined Growth Medium can use Endothelial Cell Growth Medium 2 (C-22011). In this medium, ECGS is replaced by VEGF, IGF, and additional bFGF and EGF to stimulate endothelial cell growth.