Yes, we do discourage attachments. Links are better.
This is a strong recommendation rather than a hanging offence (we are
easy-going people).
I've added a sentence to that effect at http://www.ccp4.ac.uk/ccp4bb.php
Regards
Martyn
On Thu, 2008-01-31 at 08:58 +0000, Frank von Delft wrote:
> Actually, it's not book keeping, it's simple courtesy -- and not only on
> a BB: an attachment is lazy, and a large attachment is downright rude.
>
> I am routinely stuck with a slow connection (travelling), and others are
> *permanently* stuck with one. So please be nice... ;)
>
> phx.
>
>
>
> Anastassis Perrakis wrote:
> > Dear all -
> >
> > Sorry to intervene on a 'book keeping' issue, but indeed over the last
> > few months an increasing number of people (Jerry is not the first, so
> > Jerry please do not take it personally) attach pictures etc. I think
> > in a bb standard practice dictates to only use text - if illustrations
> > are needed to explain the problem, you can put them in eg a web site.
> >
> > Some text like that was in the 'code of conduct' off ccp4bb in the
> > past, but I could no longer find it.
> >
> > Thus apologies if I am wrong and policies have changed, but maybe the
> > ccp4 crowd could tell us what is the suggested policy.
> >
> > And, if you really want to send an image please do bother to make it
> > small. The initial posting had a 630k image, which it took me 1 min to
> > make 20k and it still makes the point (attached so I can also violate
> > the rules i am suggesting - I love inconsistency).
> >
> > Thanks, Tassos
> >
> >
> >
> >
> > On Jan 30, 2008, at 20:11, Jerry McCully wrote:
> >
> >>
> >> Dear All:
> >>
> >> Thanks a lot for the prompt reply on this topic of salt
> >> sensitive complex.
> >>
> >> Attached please find one ITC final figure done under 25mM
> >> Tris(pH8.0), 60mM NaCl.
> >>
> >> As mentioned before, the ionization of Tris will interfere with
> >> the ITC experiments.
> >>
> >> Therefore I am sure of my binding results.
> >>
> >> Can anyone give me some comments on this ITC experiment?
> >> Basically do these two proteins bind to each other? If so, how should
> >> I improve the ITC experiments to get a similar affinity shown by
> >> BIAcore(about 0.5uM)?
> >>
> >> Thanks again.
> >>
> >> Jerry
> >>
> >> ------------------------------------------------------------------------
> >>
> >> Hi Jerry,
> >>
> >> Tris can cause problems, you are better off using something like
> >> HEPES, and HEPES should be ok at pH 8. (Buffers with an
> >> ethane-sulphonic acid group tend to be the best - those ending in
> >> 'ES', so MES, TES and HEPES)
> >>
> >> FYI, the error on your K is bigger than the actual measurement -
> >> 1.49x10^5 ± 1.5x10^5.
> >> Signal to noise to is probably your enemy, which is making the curve
> >> fitting difficult. Changing buffer may help this - there may be some
> >> non-specific component to what you're seeing - increasing salt a bit
> >> or dropping in something like 5% glycerol may help with this.
> >>
> >> Would you be able to post a jpeg/pdf of the curve?
> >>
> >> Regards,
> >>
> >> David
> >>
> >>
> >> On 25/01/2008, Jerry McCully wrote:
> >> >
> >> > Dear All:
> >> >
> >> > Firstly I would like to thank many folks here for giving me great
> >> > ideas several days ago.
> >> >
> >> > The following are some updates for this question.
> >> >
> >> > I did ITC experiments again using 25mMTris(pH8), 60mM NaCl(low salt
> >> > condition).
> >> >
> >> > But things still turn out to be a little weird.
> >> >
> >> > I increased the concentration of both proteins(60uM in the cell and
> >> > 1200uM in the syringe). At the end of the ITC, I saw a little of
> >> > precipitation of both the proteins.
> >> >
> >> > Fortunately I can roughly fit the curve this time. However, the heat was
> >> > still low, around 1Kcal/mole of per injectant. I am not sure about the
> >> > fitting statistics.
> >> >
> >> >
> >> >
> >> > N 1.10 ±0.17
> >> >
> >> > K 1.49E5 ±1.5E5
> >> >
> >> > DH -893.5 ±213
> >> >
> >> > DS 20.7
> >> >
> >> > Was the enthalpy was offset by the ionization of Tris buffer?
> >> >
> >> > Can I use Hepes buffer around pH8 to do ITC?
> >> >
> >> >
> >> > Welcome any comments about the statistics and suggestions on how to
> >> > improve the ITC experiments.have a nice weekend.
> >> >
> >> > Jerry
> >> >
> >>
> >>
> >>
> >> ------------------------------------------------------------------------
> >> Helping your favorite cause is as easy as instant messaging. You
> >> IM, we give. Learn more.
> >> <http://im.live.com/Messenger/IM/Home/?source=text_hotmail_join>
> >>
> >>
> >> ------------------------------------------------------------------------
> >> Shed those extra pounds with MSN and The Biggest Loser! Learn more.
> >> <http://biggestloser.msn.com/>
> >> <test-ITC-012608.JPG>
> >
>
--
***********************************************************************
* *
* Dr. Martyn Winn *
* *
* STFC Daresbury Laboratory, Daresbury, Warrington, WA4 4AD, U.K. *
* Tel: +44 1925 603455 E-mail: [log in to unmask] *
* Fax: +44 1925 603825 Skype name: martyn.winn *
* URL: http://www.ccp4.ac.uk/martyn/ *
***********************************************************************
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