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Protein concentration and extinction coefficient

Protein concentration Extinction coefficient

Best Answer mdfenko, 14 January 2014 - 04:57 AM

with lowry-type protein assays, you're reading a common colored reaction product, not the protein itself.

 

unless you empirically determine a different response of the reagent to your protein then using bsa is perfectly acceptible, with no correction factor for extinction coefficient necessary.

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#1 nitrox

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Posted 13 January 2014 - 05:40 PM

Hi all,

 

I have a question regarding estimation of protein concentration for a recombinant protein. I am using Bio-Rad DC reagent, which measures absorbance at 595 nm. To obtain protein concentration, I use a standard curve generated using BSA protein standard. Typically, I dilute my protein 2000 fold to obtain the concentration within the range. Here's my question:

 

If my protein's extinction coefficient is 22000 and for the estimation of its concentration, I have used BSA as a standard (extinction coefficient = 44000), should I double the value obtained for concentration in mg /ml, for accurate estimation? 

 

Thanks in advance! 



#2 mdfenko

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Posted 14 January 2014 - 04:57 AM   Best Answer

with lowry-type protein assays, you're reading a common colored reaction product, not the protein itself.

 

unless you empirically determine a different response of the reagent to your protein then using bsa is perfectly acceptible, with no correction factor for extinction coefficient necessary.


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#3 nitrox

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Posted 16 January 2014 - 07:18 AM

That makes sense. Thanks mdfenko! 







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