Technical library

The difference between PromoCell NHEK and NHEK GM3 lies in the growth medium used during cell isolation:
NHEK (Normal Human Epidermal Keratinocytes) are isolated in Keratinocyte Growth Medium 2 (C‑20011), a serum‑free medium that contains bovine pituitary extract (BPE).
NHEK GM3 are isolated in Keratinocyte Growth Medium 3 (C‑20021), an improved serum‑free and BPE‑free medium, offering a more defined culture environment.
 
Both NHEK and NHEK GM3:

• are isolated from the epidermis of juvenile foreskin or adult skin, and
• are available from single‑donor or pooled‑donor preparations.

Yes. Normal Human Epidermal Keratinocytes isolated in Keratinocyte Growth Medium 2 (NHEK‑GM2; C‑12001, C‑12003, C‑12005, C‑12006) also grow in PromoCell Keratinocyte Growth Medium 3 (C‑20021).
When cultured using the protocol with fixed medium‑change intervals, these cells:

• grow slightly faster than in Keratinocyte Growth Medium 2, and
• achieve more than 15 population doublings (PD).

Conversely, NHEK isolated in Keratinocyte Growth Medium 3 (NHEK‑GM3; C‑12011, C‑12013, C‑12015, C‑12016) can also be cultured in Keratinocyte Growth Medium 2 (C‑20011).
When using the classical subcultivation protocol (subculture at 70–90% confluency), these cells:

• grow slightly slower than in Keratinocyte Growth Medium 3, but
• also reach more than 15 population doublings.

No. Frozen PBMCs are not suitable as starting material for dendritic cell (DC) generation using PromoCell DC Generation Medium (C-28050).
Peripheral blood mononuclear cells (PBMCs) consist mainly of lymphocytes and monocytes. During cryopreservation, CD14⁺ monocytes significantly lose their ability to attach to tissue culture plastic. As a result, the monocyte purification step using Monocyte Attachment Medium does not work with frozen PBMCs.
 
Recommended alternatives
You can generate dendritic cells by starting with:

• Cryopreserved CD14⁺ monocytes (C‑12909), or
• Freshly isolated mononuclear cells, or
• Fresh CD14⁺ monocytes

Please refer to the Instruction Manual for DC Generation Medium (C‑28050) or the Application Note “Generation of monocyte‑derived Dendritic Cells” for detailed protocols.

This depends on the specific airway epithelial cell type.

HNEpC and HTEpC
Yes, these cell types are compatible with PromoCell ALI Medium.
Please refer to the Application Note "Air-Liquid Interface Culture of Nasal and Tracheal Airway Epithelial Cells" that specifically describes the differentiation of HNEpC and HTEpC at the air-liquid interface (ALI). It provides the recommended conditions for successful ALI differentiation of these cell types.
HSAEpC
No. PromoCell ALI Medium is not suitable for differentiation of HSAEpC at the air-liquid interface.
Therefore, no ALI differentiation protocol is provided for this cell type.
HBEpC
For Human Bronchial Epithelial Cells (HBEpC), PromoCell offers special ALI pre‑screened lots.
These lots have been successfully tested for barrier function as part of PromoCell’s quality control. For availability and detailed information on ALI pre‑screened HBEpC lots, please contact PromoCell Scientific Support.

PromoCell primary cells are cryopreserved using a computer‑controlled freezing process in the gas phase of liquid nitrogen and subsequently stored in the liquid phase of LN₂.
Cryopreservation in liquid nitrogen is a well‑established method for long‑term storage of primary cells and stem cells. When continuously stored in liquid nitrogen, PromoCell cells can be maintained for more than 10 years without loss of viability.
 
Supporting evidence
Long‑term stability of cryopreserved primary cells has been demonstrated in the literature. For example, Kumar et al. showed that adipose‑derived stem cells stored in liquid nitrogen for approximately 12 years retained viability, stem cell characteristics, differentiation capacity, and regenerative potential.

No. PromoCell has not tested the cultivation of assay‑ready M1 macrophages in MEM α supplemented with FBS.
Because PromoCell’s M1 macrophage system is fundamentally different from MEM‑based media, we cannot predict whether cell viability, phenotype, or function would be maintained under these conditions.
For reliable performance, we strongly recommend using PromoCell M1 Macrophage Generation Medium XF in combination with fibronectin‑coated culture vessels, as this is the only condition validated by PromoCell.

Yes. Below is a short, standardized protocol for expanding HPCs using HPC Expansion Medium XF supplemented with Cytokine Mix E.
 
Short protocol
1. Thawing cryopreserved HPCs
Thaw the cells for 2 minutes at 37°C in a water bath.
Transfer the cells into 9 ml complete HPC Expansion Medium XF (supplemented with Cytokine Mix E) and determine the cell count.
2. Initial seeding
Centrifuge for 10 minutes at 240×g.
Aspirate the supernatant and resuspend the pellet at 10,000 cells/ml in complete HPC Expansion Medium XF.
Seed the cells into an appropriate suspension culture vessel (e.g. 20,000 cells in 2 ml cell suspension per well of a 6‑well plate) and incubate for 2–3 days at 37 °C and 5% CO₂.
Alternatively, plate freshly isolated HPCs at a density of 10,000 cells/ml in cytokine-supplemented complete medium in a suitable suspension culture vessel.
3. Cell expansion 
After 2-3 days, add fresh complete medium to double the culture volume (e.g. add 4 ml fresh medium to 4 ml culture, resulting in a total volume of 8 ml).
Continue incubation for 10-12 days.
Perform a partial medium change every 2-3 days
Example: If the culture volume is 8 ml, discard 4 ml after centrifugation; resuspend the cells in the remaining 4 ml, then add to 12 ml fresh complete medium (final 16 ml). 
5. Harvest expanded HPCs (day 13-15)
 
For more details, please refer to the corresponding Application Note.

Yes. PromoCell Normal Human Epidermal Melanocytes derived from juvenile foreskin (NHEM.f, C‑12400) can be cultured in PromoCell Melanocyte Growth Medium M3 (C‑24310).
Under these conditions, NHEM.f can achieve more than 15 population doublings, provided that the recommended culture conditions are followed.

PromoCell recommends using Human Serum AB, off‑the‑clot, for the generation of M0 macrophages.

Osteogenic differentiation of human mesenchymal stem cells (hMSC) using PromoCell MSC Osteogenic Differentiation Medium typically takes approximately 14 days.
A detailed, validated protocol is provided in the Product Manual and the corresponding Application Note.
 
Recommended culture conditions

• Use collagen‑ or fibronectin‑coated tissue culture plates.
• Change the medium every third day during differentiation.

Maintenance after differentiation
After approximately 2 weeks, when osteogenic differentiation is complete, the differentiated bone cells tend to detach from uncoated plastic surfaces.
For this reason:

• Functional assays and analyses should be performed promptly after differentiation.
• Cells should be maintained in MSC Osteogenic Differentiation Medium until the end of the experiment.