Most primary cells detach within 5-10 min at 37°C. Inactivation isn't required but we recommend to centrifuge the cell suspension to remove accutase and EDTA before replating the cells.

Yes the specialized PromoCell media already contain the optimal amount of L-glutamine. Please don't add extra L-glutamine as this can be toxic for the cells.

PromoCell HUVECs are freshly isolated from umbilical veins. They are cryopreserved at the end of primary culture. After revival; they can be propagated for at least 15 doublings and will senesce eventually. HUV-EC-C from ATCC is a hypodiploid human cell line of endothelial origin (umbilical vein). The modal chromosome number is 45 occurring in 72% of cells counted. The rate of polyploid cells is 15.8%. The cells have a life expectancy of 50 to 60 population doublings. This indicates that the cells are no longer "normal cells" but have undergone some degree of transformation.

Our HUVEC single donor (C-12200) are isolated from a single umbilical cord; propagated in primary culture; and frozen down at subconfluency. For the preparation of HUVEC-pooled (C12203); we simultaneously isolate the cells from 2-4 umbilical cords and grow them in separate tissue culture dishes. The cells are pooled after trypsinization given that their growth rates are comparable. After thawing; our HUVECs (single donor and pooled) are both in P1. The recommended media are Endothelial Cell Growth Medium (C-22010) or Endothelial Cell Growth Medium 2 (C-22011). With respect to cell growth; HUVEC-pooled tend to have a more heterogeneous morphology with slightly more elongated cells but the doubling times are comparabel for both types (typically 18-36 hrs per doubling).

If you are growing our NHEK in PromoCell Keratinocyte Growth Medium 2 (C-20011) or Growth Medium 3 (C-20021) you don't need any feeder cells. The cells will grow as a monolayer in conventional tissue culture flasks.

Supplement Mix for Melanocyte Growth Medium M2 (C-39420) is free of serum and doesn't contain any PMA. The qualitiative and quantitative composition is proprietary.

Note: Melanocyte Growth Medium M2 and NHEM M2 cells were disontinued at the end of 2024.

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.

The time needed to detach our primary cells depends on many different factors like the cell type; cell density; lot #; trypsin concentration; the efficiency of the washing step before adding the trypsin and the trypsinization temperature. For most cell types we recommend trypsinization at room temperature and direct observation of detachment under the microscope. This way; you can find out your individual trypsinization time and keep the contact time between cells and trypsin to a minimum. Most cells detach after 2-8 min. Please refer to the instructions in the Manual. For some cell types; trypsinization at 37°C or the use of Accutase or another Detachment Solution is recommended.

PromoCell Blood and Blood Progenitor Cells are cryopreserved directly after isolation (= P0). They haven't been in culture before freezing.

We use a classification system similar but not identical to the Fitzpatrick Skin Classification. The Fitzpatrick classification has six different categories (phototypes I-VI) which correlate with the level of skin pigmentation (melanin) and sunburn following sun exposure. Fitzpatrick I corresponds with the lightest of skin complexions, while Fitzpatrick VI corresponds with the darkest skin.

I: Pale white skin, blue/hazel eyes, blond/red hair, always burns, does not tan
II: Fair skin, blue eyes, burns easily, tans poorly
III: Darker white skin, tans after initial burn
IV: Light brown skin, burns minimally, tans easily
V: Brown skin, rarely burns, tans darkly easily
VI: Dark brown or black skin, never burns, always tans darkly

At PromoCell, we have knowledge of the patients’ skin color (white, brown or black skin), color of eyes and hair, but we don't have any details about the burning/tanning abilities. We therefore classify our tissue donors as follows:

Light (comprising phototypes I and II)
Moderate (comprising phototypes III and IV)
Dark (comprising phototypes V and VI)

Information on the phototype is available for most cell lots isolated from juvenile or adult skin.

Short description: Unpack the box and place the T25 flask(s) in the incubator for 3 hrs (closed cap). Then check confluency under the microscope. When the density is < 70%; aspirate the medium using sterile conditions and add 5-10 ml of the appropriate Growth Medium. The cells should be subcultured according to the subcultivation protocol given in the cells' Instruction Manual once they have reached > 70 % confluency.

1) For fluorescence detection (fluorometer): Black plates with clear bottoms; often clear plates will suffice 2) For luminescence detection (luminometer): White/opaque plates 3) For colorimetric detection (photometer): Clear plates

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