I'm trying to detect a receptor on the surface of my A549 cells.
After trypsin treatment, I see no specific labelling although previous published report the expression of my target.
One paper stated that after trypsin treatment, they let the cell recover under agitation during 6hours in order to allow the receptor expression to come back.
as I would like to skip the incubation phase, has anyone got advice of how can I harvest my cells without trypsin?
Thanks in advance for your contribution.
Alexis
An alternative to using trypsin is Accutase which is a solution of proteolytic and collagenolytic enzymes (different from trypsin) which may not have an effect on your membrane receptor. Another option is to use a "Cell Stripper" solution which is non-enzymatic solution of chelators. Be aware when using the chelator solution that although the cells may release from the plastic, they may be "clumpy" and be difficult to disaggregate.
Good Luck!
You can also try gentle scraping of the cells to remove them from the flasks or dishes.
There are sterile cell scrapers designed for this purpose. They are very effective in removing A549s
Good luck!
Hi Alexis,
A complete alternative is to stain your cells without dettaching them and analyse fluorescence by microscopy.
At least you will know if your cells express the receptor you are looking for.
Good luck
Jacques
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