My hMSCs are failing to adhere to T-25 tissue culture flasks post-thawing.
I sterilize my culture flasks by exposing them to hood UV for one hour. Is it good practice to do so, and also does this procedure damage the treated surface?
The brand I use is BD Falcon.
Hi Shashank, there should be no need to sterilize tissue culture flasks. They are already sterilized by the manufacturer.
As for cells not adhering, I have read anecdotal reports that UV can affect TC plastic.
Anyway, skip the UV step and see it that solves the problem.
The culture flasks do not need to be exposed to UV, as they should be sterile from the company. The culture dishes should also be sterile. If you are unsure, contact your provider to ensure. Otherwise, you should be able to work with your materials without pre-exposing them to UV. In order to avoid any possible contamination, I would open all my materials in the laminar hood.
No not really, if UV was enough, autoclaves were not required. More so, the plastic petriplates are already sterilized by the company and can be kept further in the UV for some time, especially after pouring. If you are using glass petriplates then autoclave is enough to sterilize them.
The plastic flasks and petri dishes are sterilized by manufacturers. They usually use the Cobalt source of radiation. In the hood, if the distance of dishes or flasks from UV lamp is more than 30 cm, no sterilization effect could be achieved!
If you usually sterilize your tissue culture flask in UV and before you never experienced this problem then I think the cells you are using for plating after thawing are either very old, I mean you are using after many passages or you kept your cells outside for long time after taking them out from liquid nitrogen. Other thing may be the T-25 flasks you are using does not support your cells for attachment like sometime some cells attach better to tissue culture flasks if they are coated with some poly-D-lysine residues that favors attachment of cells to the surface. Well I don't think UV exposure is a problem here.
Thank you everybody for your answers ,they were really helpful. Just out of curiosity i would like to know whether
1) hood UV (generally UV-C type) has enough energy to pass through the plastic material of T-25 or any other type of cell culture flask / multi well plates.
2) if yes , can this UV light damage or modify the treated surfaces of the culture flasks on interaction with it.
3) while sterilizing the hood with UV for 15 min , we keep cell culture media , serum and trypsin inside the hood as well ( which also come in contact wih UV) . Is this a good practice . I have read that certain components of media like riboflavin and some amino acids are degraded by UV. I don't know what effects UV can have on serum or trypsin. On the whole can i suggest my peers to discontinue this practice.
If anybody has evidence for these queries in the form of research articles or write ups , i request them to kindly paste the link for the same. Thank you
It is highly discouraged to place prepared media in a culture hood with the UV light on. Hopefully you are using some sort of aseptic technique to prepare your reagents/media before and the UV light can degrade growth factors and other light sensitive agents. I would strongly encourage you to avoid doing this.
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