1. two changes of Calcium and Magnesium free Puck's Saline G (10 min ea)
2. Remove the cells with Puck's Saline G with 0.1 M EGTA.
I have gone to confocal microscopy to analyze the integrins and laminin
receptor in an attempt to not disturb the receptors.
Good luck,
Deb Berglund
Montana State University
On Mon, 25 Nov 1996
Lesterjp@aol.com wrote:
>
> We are performing surface phenotyping studies using normal and transformed
> osteoblast-like cells. Specifically, we are investigating integrin and other
> adhesion molecule expression.
>
> Thus, the obvious drawback is how to detach these cells from their substrate
> without altering surface proteins. We have tried various enzymatic
> treatments using dispase, pronase, collagenase (crude prep.), and mixtures of
> these. Cell scraping without treatment destroys the cell population. While
> dispase and collagenase are both successful in detaching the cells, I
> certainly question the maintenance of epitopic integrity following treatment.
> We have received positive (and most importantly, reproducible) results for
> some, but not all, of the mAbs tested.
>
> Any suggestions?
>
>
>