Technical library

Our HPMEC are isolated from peripheral lung tissue of adult donors.

Our HUtMEC are isolated from the middle layer of the uterine wall (myometrium). The tissue donors were not pre-treated with hormones.

Our HCMEC (Human Cardiac Microvascular Endothelial Cells) are not endocardial cells. They are isolated from the capillaries in the heart muscle. Therefore, they are in fact microvascular.

PromoCell offers two different DC Generation Media, optimized for different starting materials.
 
Freshly isolated mononuclear cells (MNC) or monocytes
For freshly isolated MNC or monocytes, PromoCell recommends DC Generation Medium XF (C‑28052). This serum‑free and xeno‑free medium is optimized for the direct differentiation of monocytes into immature and mature myeloid dendritic cells.

Cryopreserved monocytes
For cryopreserved monocytes, PromoCell recommends DC Generation Medium (C‑28050).
This medium is optimized to support dendritic cell generation after thawing of monocytes.
 
Important note for fresh MNC with DC Generation Medium (C‑28050)
If DC Generation Medium (C‑28050) is used with freshly isolated MNC, an additional initial step is required:
Monocyte Attachment Medium (C‑28051) must be used first to ensure efficient adherence and enrichment of the monocyte fraction.
 
Further details are provided in the Instruction Manual and the corresponding Application Note: Generation of monocyte-derived Dendritic Cells (moDCs).

Usually, we do not recommend specific plasticware but we have found that with PromoCell’s Dendritic Cell and Macrophage Generation Media, choice of plasticware can have a lot of an influence. For our Dendritic Cell Generation Media we recommend to use tissue culture vessels from BD Falcon^TM.

There is no significant difference in the percentage of cells that will differentiate into Dendritic Cells. But when using cryopreserved cells, the initial cell loss will be higher compared to when fresh cells are used. i.e. the final number of differentiated cells that can be expected will be higher with fresh cells as a starting material due to lower cell death rate.

It is not recommended to leave the blood cells in the Monocyte Attachment Medium for longer than 1.5-2 hrs. The medium was developed for (short-term) attachment of the monocytes and does not provide nutrients for a longer time period. Leaving the cells in Monocyte Attachment Medium for a longer time or even overnight will induce apoptosis and lead to the loss of the cells. If necessary, you can reduce the incubation time to 1 hr. In this case, it is advisable to equilibrate the media in the incubator before so that you can immediately and directly add the appropriate amount of PBMC suspension.

This method produces large numbers of adherent monocytes in only 1.5 h. If the washing steps are properly performed, 80-90% purity can be expected. The attached cells are “untouched”, since no binding of magnetic microbeads has occurred. This also excludes phagocytosis of the microbeads by the monocytes, an event which is unfavorable with regard to cellular health. In addition, the adherence method is time-saving and cost-effective.

Several different methods for the detection of mycoplasmas have been described, like e.g., cultures on agar, in liquid or semi-solid media, staining with DAPI, mycoplasma-specific antibodies, biochemical methods, and PCR-based assays. PCR-based detection is very sensitive, detects all mycoplasma species that occur in cell cultures and is completed within 3-5 hours.

No, mycoplasma can only be observed through electron microscopy. For highly sensitive detection of mycoplasma contamination, we recommend the use of PCR-based mycoplasma tests.