I have been using the EBM-2 media (along with the EGM-2 supplements) from Lonza to grow human endothelial cells and I have to say, it is perfect. However, one drawback is the price. The media is very expensive and I have been recently thinking of switching my cells to another media. I know this question has been discussed few years ago, but there seems to be no proper feedback on what happened with the researchers who asked earlier.
Integrating all what I read, I am thinking of simply going for RPMI media with 10% FBS and 1% Pen/Strep. What do you think?
Any answer is highly appreciated!
Primary cells can survive in RPMI-1640 supplemented with serum growth factors. However, their growth rate deceases over the course of few days. Further, there may be issues concerning alterations in gene expression and reproducibility of the experiments. Considering the above factors, it is not advisable to change the medium to reduce the cost of studies.
Switching to RPMI is a huge switch, which may have unwanted effects. Switching to Promocell, cheaper, yet highly similar may be an option to safe costs.
Primary freshly isolated cells have a much higher biological relevance than immortalized cell lines. If you use endothelial specific medium you also create a much more biological enviroment with defined components and growth factors. With FBS you add a cocktail of undefined growth factors and probably get different results from different FBS lots. You invest a lot of time to isolate the cells or buy them, so it is worth to invest in the defined cell culture as well.
Hi Alaa, maybe 10%FBS wont be enough, I recomend tried with 12%, also IL-2 could be necessary. You must evaluate the quality and the cost of your new media in order to establish if it is worth.
Dear Alaa,
There are good reasons why you should NOT switch to RPMI1640!
If you go back in the literature you will discover (since lost in the passage of time) that RPMI1640 was designed to support the survival and growth of cells that grow in suspension; non-adherent! To achieve this RPMI1640 is low in cations.
So what?
The so what is that endothelial cells are anchorage-dependent; they require attachment to adhesion proteins to stimulate intracellular signals that support survival. These interactions (among other requirements) are cation-dependent.
More 'so what' is that when you transfer your cells into a low cation formulation (RPMI1640) you will deprive your cells of these essential survival interactions and signals. They will likely look fine for the first/second passage, but by the time your cells have exhausted their limited intracellular cation stores they will start to die, rapidly. Many people find this confusing, failing to connect the death of their cultured cells with the changed conditions made several weeks earlier; short-term memory loss!
If you want to wean your endothelial cells onto a cheaper medium, you must also consider what else is happening! Just because your cells are still alive, does not mean they behave and respond as they would in vivo. Recall that ex vivo environments are nothing like in vivo environments and thus, cell behaviour bears no resemblance to events that take place in vivo! The question you should ask is, what is the relevance of my in vitro cell model system to the phenomenon I 'think' I am studying?
Think carefully before you change media formulations. They are created that way for a reason. If it ain't broken, it is likely to a false economy!
PS. I have no relationship with Lonza, nor any other media manufacturer. I am merely an old classical cell biologist who learned in vitro mammalian cell culture before TC Plastic was invented!
Dear Alaa, I use RPMI + 5-10% FCS + heparin + ECGS/aFGF for all of my primary EC isolations and subsequent culture (rat, rabbit, horse, cow, pig). The EC love the RPMI/ECGS combo, and the heparin keeps any SMC/fibroblasts away (as these may be present when you first isolate your cells from the vessel wall). While I've not used RPMI on human EC myself, other people in the lab do, without problems. I don't use mid or late passage or transformed EC, so cannot comment on the effect of RPMI on these cells. Good luck! PS My understanding is that EGM2 supply from Lonza will not back up until next year.
I never used anything other than EGM for endothelial cells. However, in a previous lab M199 media supplemented with
- Glutamine 0.2 M - HEPES 1 M - NaHCO3 7.5 % (w/v) - Penicilline/streptomycine (10000/10000) - FBS (heated at 56°C and centrifuged) 2% (which I find somewhat low) was used for isolating and growing HUVECs. I only tried it once - and wasn't particularly successful, but that could have had many reasons. Anyways, I switched to microvascular cells and Lonza EGM for the rest of my project.
I used RPMI 1640 for human bladder cancer cells, which are a lot less delicate.
I am always one for being economical, and I know that companies try to sell you their expensive specialised media, and sometimes it pays off to have a look around but research is expensive. My time is more valuable than testing the cheapest possible media.Perhaps you can save money in other places :).
Hi, I have HUVECs and I need to co-cultivate them for 24 hours with transfected THP-01, I just tested the survival of HUVECs in cca 20hrs in classic RPMI(+FBS,+Atb) and they seem ok, so I guess RPMI is not a problem for a short-time cultivation, but for keeping my culture growing and satisfied I use the endothelial medium.
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