Have a question about one of our products? Check out our technical library for recently asked questions from other scientists around the world.
Technical Library
Recent Entries in Technical Library
290 Entries on 29 Pages
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Can PromoCell supply preadipocytes (subcutaneous, visceral) from donors with obese BMI / non-smokers / non diabetics? Can PromoCell supply preadipocytes from different age groups?
The basic information we receive from the surgeons about the tissue donors usually includes age, gender, and ethnicity.
- For many of our donors of subcutaneous preadipocytes, we also have information on BMI, hair color, skin pigmentation, and, in some cases, smoking habits or known diseases (e.g. Diabetes). Most of our subcutaneous HWP donors are between ∼25-65 years old.
- The visceral HWP donors are mostly between ∼20-75 years old. For many cell lots we know the BMI, in some cases also the hair color, skin pigmentation, smoking habits, and/or known diseases (e.g. Diabetes or COPD).
If you are looking for particular specifications, please contact our Technical Customer Support, so that we can offer you appropriate HWP lots.
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Which type of endothelial cell is best suited for studying angiogenesis?
The study of angiogenesis has been significantly advanced by the ability to culture endothelial cells in vitro. Initially, large vessel ECs, such as those isolated from the human umbilical vein (HUVEC) were used for these studies but increasingly it has been recognized that microvascular endothelial cells are a more appropriate model since angiogenesis involves microvessels rather than large vessel ECs.
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If I isolate fresh CD14-monocytes for M1/M2 macrophage generation, can I culture them for a period of time before differentiation?
This is not advised. Please seed the freshly isolated CD14-monocytes immediately in the Monocyte Attachment Medium. Adding a culturing step will change the biological characteristics of monocytes very rapidly.
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Can the medium for M2 macrophages be switched into RPMI or RPMI + M-CSF after differentiation?
Unfortunately, we did not test this in our hands, and it must be tested by the customer. In fact, our medium is completely different from RPMI and therefore we cannot predict if this is working. We only know the successful long-term culture from our system with our media.
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Are there any differences in the cultivation protocol when cultivating MSCs compared to other cell types?
Yes, there are a few differences:
– We recommend replacing the MSC Growth Medium XF (C-28019) 3-4 h after seeding, as opposed to 16-24 hours after seeding for most other cell types/growth media, including MSC Growth Medium 2.
– When MSC Growth Medium XF, MSC Neurogenic Differentiation Medium, MSC Adipogenic Differentiation Medium 2 or MSC Osteogenic Differentiation Medium are used, flasks have to be coated with 10 µg/cm² (human or bovine) fibronectin according to the instruction manual.
– We strongly recommend using Accutase (C-41310) for cell detachment instead of Trypsin. If Trypsin is used, contact time should not exceed 2 min.Related Links and Documents
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Can I use a serum-free medium as negative control in the chondrogenic MSC differentiation? I would like to replace the recommended DMEM+10% FCS.
You could probably use our MSC Chondrogenic Differentiation Medium without inducers (C-28014). It is also serum-free and the same as C-28012, just without chondrogenic inducers.
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What is the number of primary cells per vial frozen at PromoCell? How can I calculate the number of viable cells and how should I calculate the optimal plating density?
At PromoCell we guarantee for our primary human cells ≥ 500,000 viable cells after thawing. For this, we dispense > 500,000 cells per cryovial before cryopreservation as there will always be a certain percentage of dead cells after freeze/thaw.
In order to know the number of cells that survived the procedure, we defrost a representative number of vials per lot during QC, determine the cell viability using an electronic counting device and then calculate the number of viable cells that can be recovered after thawing. Both numbers – the calculated number of viable cells and the viability – can be found on the lot-specific Certificate of Analysis (CoA) that can be downloaded from our website.
Example: When the CoA indicates 600,000 viable cells and a viability of 80%, this means that the vial actually contains 750,000 cells (viable + dead), 80% thereof (600,000) were viable after thawing in our QC. We do not indicate the total number of cells per vial but just the number of expected viable cells which can be recovered when the recommended thawing protocol is used. You don’t have to calculate any viabilities by yourself.
When the recommended plating density for your cell type is 5,000 – 10,000 cells/cm², then the 600,000 viable cells can be plated e.g. in a T75 (corresponding to 8,000 cells/cm²) or in a T75 + a T25 (corresponding to 6,000 cells/cm²).Related Links and Documents
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I am having problems isolating RNA from peripheral blood MNC pellets. Is there any reason you can think of why this would be?
Problems in obtaining RNA with good yield and purity from mononuclear cells (hMNCs) are quite common. The reason for this is the large amount of free genomic DNA usually contained in MNC preparations. This DNA originates mostly from granulocytes which underwent lysis during the isolation of the MNC. The granulocytes are gone in the final MNC preparation, but their genomic DNA – released during cellular lysis – is still there "sticking" to the MNCs. Solution: Remove DNA prior to RNA purification by a DNase digestion step. Most commercial systems include the option for such a DNase digest.
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What is the exact localization of PromoCell’s HRCEpC? Are the cells isolated from proximal or distal tubuli?
Our Human Renal Cortical Epithelial Cells (C-12660) are isolated from the cortex of the human kidney. The renal cortex is the outer portion of the kidney. It contains the renal corpuscles, the proximal and distal convoluted tubules, and the cortical collecting ducts.
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How does PromoCell determine the phototype of skin tissue donors?
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.
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