[RASMB] dn/dc

[Dan Some]

One more thing to check: does your protein absorb at the LS laser wavelength? This is not uncommon when a metal ion is incorporated. In that case, if you are using a Wyatt detector and ASTRA software, you can apply the correction algorithm for absorbing samples.

A small protein should not be an issue. MALS regularly measures sub-10 kDa and even sub-1 kDa. Insulin measurements are spot on.

With best regards,
Dan Some
Wyatt Technology Corp.
On Mar 18, 2016 4:09 AM, Kevin Jackson <wyattuk at runbox.com> wrote:
Oh...and having the correct mass recovery does not necessarily mean the MALS
has been recently calibrated....but does show the concentration response is
probably correct...and thus that the dn/dc has to be close to the correct
value (assuming you are using DRI as a concentration detector).


Kevin





Dr Kevin Jackson
Technical Director
Wyatt Technology UK Ltd
Tel: 01440 705229
Mob: 07747038083
Skype: wyattuk





-----Original Message-----
From: RASMB [mailto:rasmb-bounces at list.rasmb.org] On Behalf Of Mattia Rocco
Sent: 18 March 2016 10:32
To: Stephen McLaughlin; Andrew Leech
Cc: rasmb at rasmb.org
Subject: Re: [RASMB] dn/dc


.... and the MW should not be constant along the peak, I forgot to add, if
oligomerization is going on....


-----Original Message-----
From: Stephen McLaughlin [mailto:stephenm at mrc-lmb.cam.ac.uk]
Sent: Fri 3/18/2016 10:55 AM
To: Mattia Rocco; Andrew Leech
Cc: jphilo at mailway.com; rasmb at rasmb.org
Subject: Re: [RASMB] dn/dc

Dear Mattia, Andrew and John,

The BSA standard recovery is as expected.  Hence, given that the Zn ions are
not going to effect the dn/dc significantly, I think you are all right in
suggesting that the error will be a compound of inaccuracy in dn/dc
calculation coupled with lower light scattering for a small protein giving
rise to an larger error that would be expected when analysing much larger
proteins.

Many thanks

Stephen


On 17 Mar 2016, at 16:08, Mattia Rocco wrote:

> I agree with John regarding the dn/dc, but maybe the absolute calibration
of the instrument could also be an issue... or the on-line concentration
determination... as Andrew Leech pointed out this morning, do you recover
the approximately correct molecular weight for, say, BSA?
>
> Mattia
>
> ________________________________
>
> From: RASMB on behalf of John Philo
> Sent: Thu 3/17/2016 4:22 PM
> To: 'Stephen McLaughlin'; rasmb at rasmb.org
> Subject: Re: [RASMB] dn/dc
>
>
>
> Stephen, I think it is unlikely the dn/dc for your protein differs by more
than 5% from your calculation. If there is a "major" discrepancy then
calibration is not the issue.
>
> John
>
>
>
> -----Original Message-----
> From: RASMB [mailto:rasmb-bounces at list.rasmb.org] On Behalf Of Stephen
McLaughlin
> Sent: Thursday, March 17, 2016 5:08 AM
> To: Mattia Rocco <mattia.rocco at hsanmartino.it>
> Cc: rasmb at rasmb.org
> Subject: Re: [RASMB] dn/dc
>
>
>
> Dear Mattia,
>
>
>
> at the minute, it would be good to know if the dn/dc increases or
decreases  and roughy how much. There is a significant discrepancy between
the molar mass measured by SEC-MALS and the expected mass from mass-spec.
>
>
>
> Best
>
>
>
> Stephen
>
>
>
>
>
>
>
> On 17 Mar 2016, at 09:53, Mattia Rocco wrote:
>
>
>
>> Dear Stephen,
>
>>
>
>> it likely will. But calculating a dn/dc is not that reliable anyway...
and what's the level of precision you need, after all?
>
>>
>
>> Best - Mattia
>
>>
>
>> ________________________________
>
>>
>
>> From: RASMB on behalf of Stephen McLaughlin
>
>> Sent: Thu 3/17/2016 10:40 AM
>
>> To: rasmb at rasmb.org <mailto:rasmb at rasmb.org>
>
>> Subject: [RASMB] dn/dc
>
>>
>
>>
>
>> Dear all,
>
>>
>
>> I have a small three fingered Zn binding protein.  Will the presence of
Zn ions affect the refractive increment that can be caluculated from
sequence?
>
>>
>
>>
>
>> Best
>
>>
>
>> Stephen
>
>>
>
>>
>
>> ______________________________
>
>>
>
>> Stephen McLaughlin, Ph.D.
>
>>
>
>> Biophysics
>
>> MRC Laboratory of Molecular Biology
>
>> Francis Crick Avenue
>
>> Cambridge Biomedical Campus
>
>> Cambridge, CB2 0QH
>
>>
>
>> Tel.: +44 (0) 1223 267891 (office)
>
>>
>
>> Email:stephenm at mrc-lmb.cam.ac.uk
>
>> _____________________________
>
>>
>
>>
>
>>
>
>>
>
>>
>
>>
>
>>
>
>
>
> _______________________________________________
>
> RASMB mailing list
>
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>
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