[RASMB] SPR query about CFCA applications

Tue Jan 14 10:10:07 PST 2014

Hi John

I have done SV on many IgG samples. 1.5 if a fair average for f/f0 - but IgG's are (mostly) flexible structures, their level of glycosylation is indeed variable, so for choice - if the exact value matters to you - it's best to measure it.

Kind regards

Arthur

Arthur J Rowe
125 Uplands Road
Oadby
Leicester
LE2 4NW

Tel: +44 116 2714502
arthur.rowe at connectfree.co.uk

On 14 Jan 2014, at 17:38, John Sumida wrote:

> Natalia and Mattia,
>  
> Thank you for your responses. I gather that in general these values are not measured for the CFCA analysis and that perhaps there is not sufficient reason to do so given the precision of the application?  I wonder how variably glycosylated proteins would affect the accuracy of this technique?
>  
> Thank you both again for taking the time to respond.
>  
>  
> Best regards,
> John Sumida, Ph.D.
> Analytical Biopharmacy Core Facility
> University of Washington
> Molecular Engineering & Sciences Institute, G22
> 3946 West Stevens Way NE
> Seattle WA 98195-1651
>  
>  
>  
> From: Mattia Rocco [mailto:mattia.rocco at hsanmartino.it] 
> Sent: Tuesday, January 14, 2014 1:17 AM
> To: Markova, Natalia (GE Healthcare); John Sumida; rasmb at rasmb.org
> Subject: Re: [RASMB] SPR query about CFCA applications
>  
> 
> The f/f0 Natalia mentions is for almost spherical shape.... if you are dealing with an antibody, then the 1.5 value might be more appropriate (but I haven't done any calculations on it... ;-) )
> 
> Best - Mattia
> 
> At 08:13 AM 1/14/2014 +0000, Markova, Natalia (GE Healthcare) wrote:
> 
> Content-Language: en-US
> Content-Type: multipart/alternative;
>         boundary="_000_41C5780FD432C74DB68C588BE66E34F614EF6B76BUDURBPA08e2kad_"
> 
> Hi John,
> 
>  
> 
> We use as default f/f0 factor for globular proteins, which is 1.2 (not 1.5). However, user can freely enter any value for this factor in Web diffusion coefficient calculator (https://www.biacore.com/lifesciences/Application_Support/online_support/Diffusion_Coefficient_Calculator/index.html?section=lifesciences&realsection=lifesciences)
> 
> You need to be registered on the Biacore website to access it.
> 
>  
> 
> Best regards,
> 
>  
> 
> Natalia
> 
>  
> 
> Natalia Markova (PhD) 
> 
> GE Healthcare
> 
> R&D Life Sciences
> 
> Rapsgatan 23
> 
> 751 84 Uppsala
> 
> Sweden
> 
>  
> 
> M  +46 727 33 58 80
> 
> E   natalia.markova at ge.com
> 
>  
> 
> www.biacore.com
> www.microcal.com
> 
> 
> 
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> From: rasmb-bounces at list.rasmb.org [mailto:rasmb-bounces at list.rasmb.org] On Behalf Of John Sumida
> Sent: den 14 januari 2014 00:20
> To: rasmb at rasmb.org
> Subject: [RASMB] SPR query about CFCA applications
> 
>  
> 
> Dear RASMB,
> 
>  
> 
> This question is in regards to an SPR application.  We are currently investigating the use calibration free concentration analysis (CFCA) to monitor the stability of macromolecular preps over time in terms of their activity.  Input parameters for this analysis are ff0, viscosity, MW and D and we are using AUC to better determine both.  Current applications seem to assume ff0=1.5, which may not be correct for monoclonal preps based on my reading of the literature.
> 
>  
> 
> My query.
> 
> 1.       Is it currently the common practice to measure these quantities and if so what is the preferred method to do so?
> 
>  
> 
> Thank you all and Happy New Year,
> 
>  
> 
> John Sumida, Ph.D.
> 
> Analytical Biopharmacy Core Facility
> 
> University of Washington
> 
> Molecular Engineering & Sciences Institute, G22
> 
> 3946 West Stevens Way NE
> 
> Seattle WA 98195-1653
> 
>  
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