Mi Sun Kang wrote:
> I culture MDCK cell to research toxicity mechaism of heavy metal.
> MDCK cell is kidney epithelial cell and always had grown very quickly in my
> lab. Because it bind to each other by tight junction, gap junction,
> desmosome and focal adhesion, it looks like very big aggregates. But
> Shape of mdck cell changed into fibroblastic and size of the cell looks very
> small. All the worse, after trypsinizing to subculture, only half the cell
> survive. Doubling time of the cell is very long.
> I want to knowf what happen to my cell!!
> Could character of MDCK cell be changed by continuous subculture?
Stephen Knight's response to your post is worth reading and taking to heart.
I'd like to expand on his thoughts a bit.
First of all you should consider who or where you got your MDCK cells from
and the possibility that you did not receive what you were hoping for. Many
a lab has been sent samples with good intentions only to discover that the
wrong reagent or a contaminated reagent was sent. If there is any doubt in
your mind I would suggest getting a good culture from a trusted source. In
the United States many of us depend on the American Type Culture Collection
(ATCC--they have a web site at http://www.atcc.org/). I do not know a source
in Korea, but I do know that a similar collection of cell lines exists in
Japan. E-mail ATCC and they can probably direct you to a contact there. The
catalog number for MDCK cells in the ATCC catalog is ATCC CCL 34.
Second, you should consider what you may have done to the cells in your lab.
The point is well made by Stephen Knight concerning cross-contamination,
mycobacterial contamination, and source contamination in your reagents and/or
laboratory ware. As a rule of thumb you should always freeze ampules of a
cell line prior to commencing experiments. It is also a good idea to pick a
passage number within which you will work. You ask in your message if the
character of MDCK cells can change with continuous subculture. The answer is
that ANY cell line can and probably will change under these conditions.
Remember that every day is evolution day in the culture flask. You are
continuously selecting for cells that grow best under your conditions.
Experiments performed on passage 100 cells may very well not replicate
experiments performed on passage 10 cells. Its an inherent problem we all
accept, but it doesn't hurt to remind oneself of this point every so often
:o) Remember that the MDCK cell line was generated in 1958. Today it is
well known that there are high resistance (type I) and low resistance (type
II) MDCK cells. One need look no further than the work performed on the
sorting of the Na+,K+-ATPase to see how difference in clones of MDCK cells
can yield contrasting results (For a good read of science policing itself see
the discussion that takes place in "Science" volume 260 pp.550--556.)
In conclusion, problems such as those you have described can be a real
nightmare. Go back to your frozen stocks if you have them and double check
your ability to replicate your data so far. Do not destroy your sick culture
until you have thoroughly tested for mycoplasma or other contamination. If
present, thoroughly decontaminate your culture area before thawing your new
cells! In addition, to be safe I would also order some bona fide MDCK cells
and check the ability to replicate your data in these as well. To safe-guard
your lab in the future try to quarantine all new cultures you get from
colleagues and test for contamination before admitting into your stock.
Best of luck,
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