A curious result- hide and seek with my protein - (Oct/06/2008 )
the other one gave no signal or poor signal?
if it gave poor signal then you should be able to optimize the signal (by adjusting the antibody concentration, reaction time, etc).
-mdfenko-
QUOTE (mdfenko @ Oct 31 2008, 08:46 PM)
the other one gave no signal or poor signal?
if it gave poor signal then you should be able to optimize the signal (by adjusting the antibody concentration, reaction time, etc).
if it gave poor signal then you should be able to optimize the signal (by adjusting the antibody concentration, reaction time, etc).
pretty poor, in your opinion there is still hope that I can detect the protein. thanks for your support tiffy
-Tiffy-
Did you do IHC with the other cell line (the one can be detected with IB)? What is the result comparing to your cell?
I still think that it is possible that your cell does not have enough protein for IB detection. Maybe concentrate the protein more, that is: separate out the portion of lysate (membrane or cytosol) that your protein expressed in and try again. I have known some friend has to run a whole 15-cm plate worth of lysate in one lane to detect a protein by IB although he could detect it in IF.
-Almasy-
QUOTE (Almasy @ Nov 1 2008, 01:52 PM)
Did you do IHC with the other cell line (the one can be detected with IB)? What is the result comparing to your cell?
I still think that it is possible that your cell does not have enough protein for IB detection. Maybe concentrate the protein more, that is: separate out the portion of lysate (membrane or cytosol) that your protein expressed in and try again. I have known some friend has to run a whole 15-cm plate worth of lysate in one lane to detect a protein by IB although he could detect it in IF.
I still think that it is possible that your cell does not have enough protein for IB detection. Maybe concentrate the protein more, that is: separate out the portion of lysate (membrane or cytosol) that your protein expressed in and try again. I have known some friend has to run a whole 15-cm plate worth of lysate in one lane to detect a protein by IB although he could detect it in IF.
Well, to be honest I haven`t never carried out an IF simultaneously with both cell lines. So I can`t say if there is a difference in the signal intensity. This would be the next step, seed cells (same cell density) and run an IF.
Tiffy
-Tiffy-