° ¹Ì ¼± 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?
>> please let me know.
> thank you in advance.
>> email to oasis at cau.ac.kr>> Mi Sun Kang
> Department of Environmental and Health Chemisty,
> College of Pharmacy,Chung-Ang university,seoul, south korea
I have not had that much experience with the MDCK cell line but
have worked with a number of other cell lines and primary cultures over
the years. I'm not aware of the MDCK spontaneously transforming in
vitro, fom epithelial to fibroblast. Have you had your cell cultures
checked for Mycoplasma/Ureaplasma contamination? The condition of your
cells makes me think that the cell line is contaminated either through
subpassaging or a component of the medium, i.e. serum. Bear in mind that
these bacteria will not be visible by routine phase contrast light
microscopy or histological/Gram stain. If your cells are not
contaminated with bacteria or virus then there may be a cytotoxic
component in your culture conditions, i.e. medium, plastics, water,
glassware etc. which will be difficult to identify or never will be. An
additional but less likely possibility is that you MDCK cell line was
cross contaminated with another cell type being used in your laboratory.
In my lab. cell cultures are routinely checked for
contamination every 6 weeks (Hoechst stain method) and discarded when
they reach 50-60 subpassages from the freezer. This strategy ensures
that there are always clean, viable cell stocks in the freezer, and that
if there are problems they can be isolated and identified. Where
possible we have specific markers (cytokeratins, receptors) for specific
cells to ensure identity and continuity.
I hope that this advice is useful and that your problem can be
Microbiology and Immunology
UNC- Chapel Hill, North Carolina