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Technical Library
Recent Entries in Technical Library
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Why are the cryopreserved macrophages abbreviated as hMDM-GMCSF(-) and hMDM-MCSF(-), respectively? What does the (-) stand for?
- hMDM-GMCSF is the abbreviation for human monocyte-derived macrophages. They are polarized [⇒ differentiated with GM-CSF] but non-activated [⇒ (-)] M1 macrophages.
- hMDM-MCSF is the abbreviation for human monocyte-derived polarized [⇒ differentiated with M-CSF] but non-activated [⇒ (-)] M2 macrophages.
The macrophages can be seeded into all kinds of TC vessels. After plating, they can be maintained as biologically functional adherent cultures for several weeks.Optionally, user-customizable activation of the cells can be performed. For details, please see attached Application Note, page 2, Fig. 3 and page 5, Tab. 1).
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I differentiated M1 macrophages from self-sourced PBMCs following your differentiating protocol. In the final cell culture, I can see small cells attached to the macrophages which are CD3+ (T-cells). What is the reason for the T-cell contamination after the differentiation?
The reason for the higher number of lymphocytes in the macrophage culture is probably due to an insufficient washing step during the purification of the monocyte via adherence. The 3 washing steps in our protocol are essential to receive a monocyte population of over 90%.
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After thawing the supplements, I see some precipitation. Is this normal?
Light flocculation may be seen upon thawing the supplements containing ECGS/heparin or BPE. This does not affect the activity of our media.
Optionally, the precipitate can be removed by centrifugation under sterile conditions.We recommend to thaw the supplements (SupplementMix or SupplementPack) at 15-25°C.
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Is it possible to thaw and re-freeze the Cytokine Mix M1 (C-39894) and M2 (C-39895)?
The Cytokine Mix M1 and M2 should not be subjected to further freeze/thaw cycles.
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Can I cultivate the macrophages in complete medium but w/o cytokines?
Without cytokines, the macrophages will die very quickly. You must supplement the Macrophage Base Medium XF (Bsal Medium + SupplementMix) with the appropriate cytokines or at least with human AB ser
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We use Macrophage Generation Media from PromoCell to get differentiated macrophages from peripheral blood monocytes. The monocytes are isolated by culturing PBMCs in Monocyte Attachment Media (also from PromoCell) for 1.5 h. Would it be ok if the attachment step is done overnight, since we receive the blood later in the day which makes it difficult to process on the same day?
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.Related Links and Documents
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Does PromoCell recommend a particular manufacturer or brand of tissue culture plastic to grow the primary cells or can I use any supplier on the market?
Our customers have successfully used TC flasks and dishes from all the leading cell culture plastic suppliers to grow PromoCell’s primary human cells. We do not have any knowledge whether the dishes from local TC plastic suppliers work in the same way. We recommend to first test whether these brands provide the same good performance as the plastic of the leading manufacturers.
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After differentiation of human MSC into mature adipocytes – what medium do I have to use to culture those cells?
Differentiation of hMSC into mature adipocytes takes approx. 2 weeks. You can keep the adipocytes for up to 3 weeks in the MSC Adipogenic Differentiation Medium. After 3 weeks we recommend to switch to PromoCell’s Adipocyte Nutrition Medium (C-27438).
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What is the composition of Melanocyte Growth Medium M2 SupplementMix (C-39420)?
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.
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What is the source of PromoCell’s Normal Human Follicle Dermal Papilla Cells? For how long can they be maintained in culture?
HFDPC are isolated from the hair papilla of normal human scalp hair follicles. Hair papilla in the adult hair follicle play a crucial role in the dermal-epidermal interactions that control hair production and in hair growth cycle events. The follicle dermal cells are cryopreserved at second passage and can be cultured for at least 10 population doublings when using PromoCell Follicle Dermal Papilla Cell Growth Medium (Cat. C-26501). Typical population doubling times are between 20-36 hrs.
The recommended seeding density after thawing/trypsinization is 5,000-10,000 cells/cm2. Using 1:4 splits, you can perform 4-5 passages with the cells.Related Links and Documents
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