I need to test whether my storage buffer will precipitate my protein. How can I know my protein is precipitated by the buffer?
0
3 replies to this topic
#2
mdfenko
-
- Active Members
-
- 2,251 posts
an elder
79
Excellent
Excellent
Posted 21 April 2009 - 08:13 AM
centrifuge it and see if it develops a pellet (before and after changing the buffer).
talent does what it can
genius does what it must
i do what i get paid to do
genius does what it must
i do what i get paid to do
#3
sharath
-
- Active Members
-
- 60 posts
Enthusiast
0
Neutral
Neutral
Posted 21 April 2009 - 01:58 PM
Treat your sample with the storage buffer and a suitable control. If the protein precipitates, a pellet will be visible upon centrifugation. However, the pellet will be discernible only if the concentration of your protein is high enough to produce a large one. In case you do not see any pellet, run SDS-PAGE with the supernatant along with the control and comapre the intensity of the bands. If you have a functional assay for your protein then try it with the supernatant. Note: do not forget to inlcude a good control; this is a compatative anlaysis.
Sharath B.
#4
Nrelo
-
- Active Members
-
- 71 posts
Enthusiast
0
Neutral
Neutral
Posted 22 April 2009 - 01:52 AM
sharath, on Apr 21 2009, 01:58 PM, said:
Treat your sample with the storage buffer and a suitable control. If the protein precipitates, a pellet will be visible upon centrifugation. However, the pellet will be discernible only if the concentration of your protein is high enough to produce a large one. In case you do not see any pellet, run SDS-PAGE with the supernatant along with the control and comapre the intensity of the bands. If you have a functional assay for your protein then try it with the supernatant. Note: do not forget to inlcude a good control; this is a compatative anlaysis.
thx guys. My protein is an enzyme so I am going to use enzyme assay for testing
