[Rasmb] Tween 80 effect on c(s)

[Qin "Chin" Zou]

Hi all,
I wonder if anyone has looked at the effect of Tween 80 on the s
distribution. I remember that John Philo once asked the similar question and
had calculated s for Tween 80 around 2s. 
I have an 18KD protein that runs at about 1.7s when there is no Tween 80 in
the buffer. However, in the presence of Tween 80, there were two species 1.2
and 1.5s (without regularization. If F=.68, then two peaks are overlapped).
The experiment was run at 60K rpm, absorbance at 276nm. Also, the fit was
not good with rmsd above 0.01. In addition, I tried to scan at 289nm, where
the absorbance of Tween 80 is minimal, with higher protein concentration
(5mg/ml). Although the 1.2s peak is not there anymore, the main peak is at
1.4s instead of the expected 1.7s. Also the fit was quite bad with rmsd
around 0.03.  For both experiments, I had 0.02% Tween in the reference
buffer.
Is it possible that the 1.2s peak is due to the Tween absorbance at the
region between the meniscuses of reference and sample sectors? I have not
tried with the same column height in both sectors. It is hard to match them
exactly. Would it be better to run this kind of experiment without including
Tween 80 in reference buffer and note the potential 2s peak as the Tween
peak? For my protein of 1.7s, it could be still difficult. Any input will be
appreciated.

Qin "Chin" Zou
Eli Lilly and Co.