Hi Amir, Ade, Mark and Tom,
Thanks a lot for your replies. Checking with BSA is a great idea and I
should've done it earlier. The BSA run for recent experiment is OK. I
cannot find a standard profile for my experiment one year ago. So I
guess that there was something wrong with the machine one year ago.
Besides, as Tom pointed out, 144KD is way off the MW of dimer. So I am
quite confident that I should use the new data.
BTW, the protein concentration is same. And it is a monomer-dimer
equilibrium (11.4ml and 10.0ml).
Zhen
Quoting Amir Khan <amirr...@tcd.ie>:
Hi Zhen
-was concentration higher in old run? Might be monomer-dimer equilibrium,
for recent paper, see Benfield et al. (JBC, in press)
-we have a miniDAWN with a 'low-high-medium' setting in the back of
the setting.
Inadvertently changing the setting scales the MW values up and down.
Have you run a BSA standard, to insure system is OK?
-Amir
________________________________________
From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] On Behalf Of zhen
zhang [zz2...@columbia.edu]
Sent: Thursday, April 21, 2011 5:32 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] Puzzle with MALS
Dear All,
I am having a consistency issue processing my SEC-MALS data. The
sample is a 80KD protein which may form a dimer. The experiment I
run a year ago shows a major peak at 11.4 ml (GE S200 column), which
is calcuated to have a MW of 144kD. There is a minor peak at 10.0
ml. In an effort to make nice looking pictures, I rerun the
experiement using the same setup (column, buffer, protein, speed)
and again the major peak is at 11.4 ml but it is calcuated to have a
MW of 80KD. I rerun the sample multiple times with the same result.
I tried to reprocess the old data and the calcuated MW is still
144KD. I am puzzled about what I have done wrong and which data I
should trust. Well, I am inclined to trust the later experiment
because I took extra caution and had multiple data sets. However, I
should be able to reprocess the old data to have a MW close to 80KD
if I can find out what is wrong and correct the mistake, which I
have not been able to. Any suggestions?
Zhen Zhang
