HI
I am trying to perform a 2D analysis with 500 ug of protein (12% PAGE). I know that the protein to SDS ratio has to be 1:1.4 Can I increase the SDS in the resolving and stacking buffers since I cant alter the amount at the sample stage? Please help. Krith
0
3 replies to this topic
#2
mdfenko
-
- Active Members
-
- 2,286 posts
an elder
83
Excellent
Excellent
Posted 28 March 2011 - 12:39 PM
you shouldn't need to increase the sds in the buffers.
have you tried running with the normal formulation? if so, what happened that makes you think you need more sds?
have you tried running with the normal formulation? if so, what happened that makes you think you need more sds?
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
krithbala
-
- Members
-
- 3 posts
member
0
Neutral
Neutral
Posted 29 March 2011 - 08:23 PM
I have tried running similar samples on 1D and found that they didnt move much. The bands remained at the top as blobs, right after the stacking gel front. I thought adding more SDS to the gel buffers would help me with the problem in 2D.
#4
mdfenko
-
- Active Members
-
- 2,286 posts
an elder
83
Excellent
Excellent
Posted 30 March 2011 - 06:55 AM
what is the expected molecular weight range of the proteins?
are you sure that 12% is not too restrictive?
have you tried a lower acrylamide concentration or a gradient?
are you sure that the protein is solubilized? did you boil in sample buffer?
are you sure that 12% is not too restrictive?
have you tried a lower acrylamide concentration or a gradient?
are you sure that the protein is solubilized? did you boil in sample 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
