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Question: Bradford assay - buffer precipitates


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#1 Benita

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Posted 18 February 2010 - 05:03 AM

Hey,

I have my protein in a 300mM KCl and 20mM Hepes Buffer pH7,9 so far. I'd tried to do a bradford but my buffer precipitates after adding solution B. I don't know what happened there? What precipitates there?

Thank you very much!

#2 mdfenko

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Posted 18 February 2010 - 08:31 AM

bradford solution b? maybe you mean lowry or bca? if that's the case then you may be seeing a precipitate due to potassium.
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#3 Benita

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Posted 04 March 2010 - 06:24 AM

bradford solution b? maybe you mean lowry or bca? if that's the case then you may be seeing a precipitate due to potassium.


I mean the second solution, used for an Bradford assay.

What kind of Protein Quantitation Assay would be possible for KCl in protein buffers?

Thank you for your help.

Edited by Benita, 04 March 2010 - 06:25 AM.


#4 mdfenko

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Posted 05 March 2010 - 12:38 PM

I mean the second solution, used for an Bradford assay.

What kind of Protein Quantitation Assay would be possible for KCl in protein buffers?

Thank you for your help.

the bradford should be okay for use with kcl. the bradford reagent can precipitate (coomassie is not as soluble in water as it is in alcohol) upon dilution. you can vortex then centrifuge to pellet any insolubles (then carefully transfer the solution to the cuvette).

when we get precipitation, due to kcl, in the lowry, we pellet by centrifugation after the incubation is complete and read the solution.
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