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<DIV dir=ltr align=left><SPAN class=351382420-26042011>Mark,</SPAN></DIV>
<DIV dir=ltr align=left><SPAN class=351382420-26042011></SPAN> </DIV>
<DIV dir=ltr align=left><SPAN class=351382420-26042011>Given that the fit is so
bad I'm not sure it makes any sense to try to get a confidence interval. The
model is clearly not even coming close to representing the data, so what does
this association constant mean? </SPAN></DIV>
<DIV dir=ltr align=left><SPAN class=351382420-26042011></SPAN> </DIV>
<DIV dir=ltr align=left><SPAN class=351382420-26042011>I agree fully with Jack
that you cannot expect the peaks from c(s) analysis to give you the monomer and
dimer sedimentation coefficients, and that you really want to do a dilution
series over a big concentration range. For absorbance data that usually means
either using multiple wavelengths (in different runs) or using the 3 mm
centerpieces to help cover a big range while keeping the OD <=1 for the high
end. Then you can either fit the weight-average sedimentation coefficient versus
concentration to a model, or do global whole boundary analysis with SEDPHAT
or SEDANAL (with the sedimentation coefficients as fitting
variables).</SPAN></DIV>
<DIV dir=ltr align=left><SPAN class=351382420-26042011></SPAN> </DIV>
<DIV dir=ltr align=left><SPAN class=351382420-26042011>For this or any other
experiment I also recommend that you fill you cells more completely to get
a better separation. Try to get the meniscus at 5.9-5.95 cm. </SPAN></DIV>
<DIV dir=ltr align=left><SPAN class=351382420-26042011></SPAN> </DIV>
<DIV dir=ltr align=left><SPAN class=351382420-26042011>John</SPAN></DIV><BR>
<DIV dir=ltr lang=en-us class=OutlookMessageHeader align=left>
<HR tabIndex=-1>
<B>From:</B> rasmb-bounces@rasmb.bbri.org [mailto:rasmb-bounces@rasmb.bbri.org]
<B>On Behalf Of </B>Mark Agacan<BR><B>Sent:</B> Tuesday, April 26, 2011 8:36
AM<BR><B>To:</B> RASMB@rasmb.bbri.org<BR><B>Subject:</B> Re: [RASMB] error
estimates and F-statistics in Sedphat<BR><BR></DIV>
<DIV></DIV>
<DIV>Thanks Patrick and Chad for the information about error estimates from
F-statistics.</DIV>
<DIV> </DIV>
<DIV>My problem now is that the apparent errors for the binding constants
derived from the analysis of F-statistic appear to be very large.
When I make a change to e.g. log10Ka12, by any value from 0.1 - 3.0, there is no
change (or only a very small change) in chi squared, so I can never reach the
critical chi squared. </DIV>
<DIV> </DIV>
<DIV>Each dataset looks good and gives low rmsd values when processing with the
c(s) model in sedfit. I'm using the monomer-dimer self-association model
in sedphat, with the values for S1 and S2 taken from c(s) analysis. I have
tried to fit to other models with only worse results.</DIV>
<DIV> </DIV>
<DIV>The fit looks very poor (see screenshots attached). I don't think I
need to repeat the experiment because the data looks good, at least to my
eye, but the fit to the monomer-dimer model seems very bad. </DIV>
<DIV> </DIV>
<DIV>Any further suggestions or advice would be very welcome.</DIV>
<DIV> </DIV>
<DIV>Many Thanks,</DIV>
<DIV> </DIV>
<DIV>Mark</DIV>
<DIV> </DIV>
<DIV><BR> </DIV>
<DIV> </DIV>
<DIV>~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~<BR>Dr Mark
Agacan, Scientific Officer for the Division of Biological Chemistry and Drug
Discovery,<BR>Wellcome Trust Biocentre, College of Life Sciences, University of
Dundee, Dundee, DD1 5EH <BR>Tel: +44 1382 386095 Fax: +44 1382
345764 Mobile: 07525 451
117<BR>~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~</DIV>
<DIV> </DIV>
<DIV>
<DIV>************************************************************ </DIV>
<DIV>
<DIV><FONT color=#666666>Please consider the environment. Do you really need to
print this email?</FONT> </DIV></DIV><BR><BR>>>> Chad Brautigam
<Chad.Brautigam@UTSouthwestern.edu> 4/23/2011 15:02 >>><BR></DIV>
<DIV
style="FONT-FAMILY: Tahoma; DIRECTION: ltr; COLOR: #000000; FONT-SIZE: 10pt">Hi,
Mark,<BR><BR>The F-statistics in SEDPHAT are meant to allow you to construct a
"do-it-yourself" error interval. Check out the statistics help page.
Under the heading "Error Analysis" there is a procedure for how to use the
output "critical chi-square" from the Statistics Menu to find an error
interval.<BR><BR>Cheers,<BR>Chad<BR><BR>
<DIV style="FONT-FAMILY: Times New Roman; COLOR: rgb(0,0,0); FONT-SIZE: 16px">
<HR tabIndex=-1>
<DIV style="DIRECTION: ltr" id=divRpF398298><FONT color=#000000 size=2
face=Tahoma><B>From:</B> rasmb-bounces@rasmb.bbri.org
[rasmb-bounces@rasmb.bbri.org] on behalf of Mark Agacan
[M.Agacan@dundee.ac.uk]<BR><B>Sent:</B> Friday, April 22, 2011 6:36
AM<BR><B>To:</B> rasmb@rasmb-email.bbri.org<BR><B>Subject:</B> [RASMB] error
estimates and F-statistics in Sedphat<BR></FONT><BR></DIV>
<DIV></DIV>
<DIV>
<DIV>Hello,</DIV>
<DIV> </DIV>
<DIV>I'm calculating errors in the binding constants in sedphat as instructed in
the statistics help page, but I don't appear to get an output for the
F-statistics. Are the F-stats output only as (changes in) chi-squared
values? e.g. the error associated with the binding constants is clearly given in
the covariance matrix.</DIV>
<DIV> </DIV>
<DIV>Best Wishes,</DIV>
<DIV> </DIV>
<DIV>Mark</DIV>
<DIV> </DIV>
<DIV> </DIV>
<DIV>~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~<BR>Dr Mark
Agacan, Scientific Officer for the Division of Biological Chemistry and Drug
Discovery,<BR>Wellcome Trust Biocentre, College of Life Sciences, University of
Dundee, Dundee, DD1 5EH <BR>Tel: +44 1382 386095 Fax: +44 1382
345764 Mobile: 07525 451
117<BR>~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~</DIV>
<DIV> </DIV>
<DIV> </DIV>
<DIV>************************************************************ </DIV>
<DIV>
<DIV><FONT color=#666666>Please consider the environment. Do you really need to
print this email?</FONT> </DIV></DIV><BR>
<P>The University of Dundee is a registered Scottish charity, No: SC015096
</P></DIV></DIV></DIV><BR>
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<FONT color=blue size=2 face=Arial><BR>UT Southwestern Medical Center<BR>The
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