HPLC Assay Question

larson eric elarson at ux1.cso.uiuc.edu
Mon Jan 30 21:27:39 EST 1995


cjhixon at netcom.com (Carl Hixon) writes:

>Secondly, I have a protein 
>suspension containing small protein particles (few microns in diameter)  
>These particles are constructed of Albumin and Fibrinogen.  Would it be 
>possible to determine the protein concentration of this type of 
>suspension using HPLC by digesting the particles?  I worry about the 
>different sizes of the protein fragments. 
==========
 
The Lowry protein procedure is likely to work fine on these samples if they
are first treated to alkaline hydrolysis by NaOH.  It's also possible that
the pH of the regular Lowry is high enough to cause dissociation of the
protein.  The Lowry also works in the presence of 1% SDS if the Cu
concentration is raised several-fold.  

Colleagues routinely pre-dissolve their protein in a small volume (10% of
final volume) of 0.1 M NaOH -- then do the Lowry as normal.  NaOH
solubilization also works after Na Deoxycholate ppt..  
 
-- 
Eric Larson                  | University of Illinois at Urbana-Champaign
USDA/Agronomy                | 190 PABL; 1201 W. Gregory; Urbana, IL 61801
elarson at ux1.cso.uiuc.edu     | Voice 217.244.3079  Fax 217.244.4419
Fidonet: 1:233/4.1           | My opinions are my own, but correct :-) 



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