Protein Concentration Estimation

Andreas Weber aweber at biolan.uni-koeln.de
Mon Jul 18 12:29:17 EST 1994


In article <gends.7.000D9E48 at leeds.ac.uk>, gends at leeds.ac.uk (SCOTT D.) 
says:
>
>Hia,
>I need an accurate method of measuring protein concentration. My sample 
is an 
>RNA binding protein, and for various reasons very difficult to get rids 
of the 
>nucleotide without also getting rid of your sample. The prescence of the 
>nucleotide is not inhibitory on the studies I'm carrying out, but it is 
>providing just a few problems in estimating how much sample is there. 
>Absorbance provides anomalous answers, and Biorad (Bradford) assays are 
out as 
>the answer needs to be accurate to within 5%. Amino Acid analysis tells 
us 
>accurately how much protein is there, but I'd love a way of doing this 
in less 
>than two days!
>So is there any ideas out there? Has anyone come across similar 
problems, 
>pointers in the right direction and information on dead ends not to 
pursue 
>would be very much appreciated!
>Cheers in advance,
>
>Dave Scott.
>Dept of Genetics.
>University of Leeds.
>U.K.
>Email: gends%south-01.novell.leeds.ac.uk at gps.leeds.ac.uk

Hi Dave,

if you have enough smple, you should load an aliquot on a SDS-PAGE
You can measure the absorption of the Comassie-stained protein
in the SDS-PAGE with laser densitometers or flat bed scanners.
Or you could do an amino acid analysis of your sample, this would
give you the most accurate values. You will need at least
50pmol of protein for correct AAA.

Hope this helps

Andreas

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