 The function of EDTA for harvest cells |
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  Jan 6 2005, 08:00 PM
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#1
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member  Group: Members Posts: 5 Joined: 6-January 05 Member No.: 5248  |
Dear all,
I am going to analyze the proteins of whole cell. The cells are cultured on surface of T-flask. usually, I use Trypsin to harvest cells. However, it is said ENTA is better since it did not destroy the surface proteins. Anyone can tell me how does EDTA work? I want to know the mechanism. Thank you!! |
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Jan 7 2005, 07:41 AM
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#2
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Enthusiast Group: Active Members Posts: 95 Joined: 13-December 04 From: Barcelona Member No.: 5058 |
EDTA acts by chelating calcium and magnesium ions from the intercellular bridges and desmosomes. Cells then dissociate from each other and from the support media. You can use 1-2% EDTA diluted in HBBS or PBS, the incubation time will depend on your cell line. EDTA is not highly efficient by itself, but potentiates the action of trypsin.
If you want to obtain intact cells you may also try scraping them. I usually trypsinize the clls for subculturing but scrape them when I'm gonna extract protein. Cheers! -------------------- Science is a wonderful thing if one does not have to earn one's living at it
(A.Einstein) |
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Jan 7 2005, 01:30 PM
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#3
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member Group: Members Posts: 5 Joined: 6-January 05 Member No.: 5248 |
Thank you for your reply!
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