Technical library

No. It is not necessary to use coated culture flasks for Nasal Epithelial Cells.
For standard cultivation, Nasal Epithelial Cells can be grown directly on tissue culture-treated plastic, and therefore coated flasks are not recommended or required in the PromoCell Instruction Manual.
 
Use of coatings in special applications

• While not required for routine culture, some customers use collagen‑coated dishes for specific applications
• Such coatings may be helpful in specialized experimental setups, depending on the research question

However, coating is not essential for normal growth and maintenance of Nasal Epithelial Cells.

Yes. Mesenchymal Stem Cell Growth Medium XF (C‑28019) does contain phenol red.
The exact concentration is confidential.
 
Phenol red‑free alternative
PromoCell also offers a phenol red‑free variant:

• Mesenchymal Stem Cell Growth Medium XF, prf (C‑28018)

This variant is suitable for applications where phenol red may interfere, such as imaging, reporter assays, or hormone‑sensitive experiments.

Difficulties in obtaining RNA with good yield and purity from hMNCs are quite common.
 
Underlying reason
The main cause is usually the presence of large amounts of free genomic DNA in hMNC preparations. This DNA originates primarily from granulocytes, which:

• Undergo lysis during the isolation process
• Are not present in the final hMNC fraction as intact cells
• But release genomic DNA that remains in the preparation and can adhere to the mononuclear cells

This residual DNA can significantly interfere with RNA isolation, leading to low purity or poor RNA yield.
 
Recommended solution
To address this issue, it is recommended to remove DNA prior to RNA purification by including a DNase digestion step.
Most commercial RNA isolation kits offer an optional DNase treatment, which is well suited for this purpose.

For optimal post‑thaw recovery, please follow the protocol below when freezing normal human cells.
Cell freezing protocol
1. Detach the cells using your standard detachment procedure.
2. Count the cells and ensure a viability greater than 70% before freezing.
3. Centrifuge the cells and resuspend them in a suitable freezing medium at a density of 0.1–10 × 10⁶ cells/ml.

• We recommend Cryo‑SFM Plus, a defined, animal‑component‑free and protein‑free cryopreservation medium.
• The freezing medium should be pre‑cooled to 4°C. Do not exceed room temperature.

4. Aliquot the cell suspension into cryovials.
5. Cool the cells slowly to −80°C at an approximate rate of −1°C per minute.

• We recommend using a controlled‑rate freezing container such as Mr. Frosty™ (Nalgene) and placing it in a −80°C freezer overnight.

6. Transfer the frozen vials to liquid nitrogen for long‑term storage.
 

Primary cells (primary culture) are freshly isolated cells that are plated in a tissue culture vessel for the first time. This initial culture stage is referred to as passage 0 (P0).
Normal cells are primary cells that have been subcultured at least once. From this point onward (i.e. passage ≥ 1), they should correctly be termed normal cells.
 
In summary, the distinction is purely based on culture history and passage number, not on cell origin or biological characteristics.
In everyday laboratory practice, however, the term “primary cells” is often used interchangeably with “normal cells".
 

Adult stem cells are stem cells isolated from postnatal tissues that have retained the capacity for self‑renewal as well as for differentiation into multiple cell lineages (multipotency).

PromoCell does not specify the number of passages that can be performed with a cell culture. Instead, we determine the number of population doublings (PDs) that can be achieved under the recommended culture conditions.
The term passage only describes the process of detaching and replating cells and does not account for differences in split ratios. As a result, the number of passages alone is not a reliable measure of a culture's expansion potential.
The optimal split ratio depends on the actual cell yield after trypsinization and the recommended seeding density. For most PromoCell cell types, typical split ratios range from 1:4 to 1:6.
As an example, when using a 1:4 split ratio (i.e. a four‑fold increase in growth surface area at each passage), approximately 15 population doublings can be achieved within 6–8 passages.
For recommended seeding densities and culture conditions, please refer to the corresponding Instruction Manual (section: Specifications).

PromoCell guarantees 15 population doublings (PD) for most normal human cells, unless otherwise stated in the Certificate of Analysis (CoA).
This guarantee applies when the cells are cultured using the recommended PromoCell growth media and PromoCell detachment reagents.

Additional information
Further details can be found in the Instruction Manual for the respective cell type under “Specifications” or on the corresponding product page in our webshop (see the “Product Description” tab).

If cells do not detach or detach very slowly during subculture, this is usually caused by suboptimal subcultivation conditions or handling.

Possible reasons and solutions

• Cells are too confluent → Trypsinize cells at a subconfluent stage
• Washing step was omitted → Perform a washing step with HepesBSS or PBS to remove residual medium before adding trypsin
• Trypsin solution is too cold → Use pre‑warmed trypsin (room temperature or 37°C)
• Trypsin solution is too old → Check the expiry date and use a fresh solution
•  Incubation time too short →  Use proper incubation time until 80% have detached 

If the issue persists, PromoCell Scientific Support will be happy to assist you in identifying the cause and optimizing your subculturing protocol.
 
Please note: Some epithelial cell types adhere very firmly and need prolonged incubation times. To avoid irreversible damage by over-trypsinization, the use of accutase is recommended. Accutase is very gentle to the cells, preserves the surface receptors and doesn’t affect the cellular viability during extended contact with the cells.

Microbial contamination is typically introduced during cell culture handling and may originate from several common sources.
 
Typical causes include:

• Unsterile working techniques (e.g. insufficient disinfection, improper aseptic handling)
• Contaminated equipment or environment such as:
  • Water in water baths
  • Incubators
  • Laminar flow hoods
• Contaminated reagents or consumables including:
  • Culture media or supplements
  • PBS, trypsin, or other solutions
  • Cell culture plasticware

Microbial contamination can severely affect cell growth and morphology.