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40mM EDTA in (non-metallo)protease assay buffer


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#1 DRT

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Posted 05 March 2014 - 03:06 PM

Does anyone know the reason for the extremely high concentrations of EDTA in casein based protease assays for papain? The AOAC/FCC/ US & European Pharmacopeia all give 14g/L (example below). To me it looks like a decimal point typo that has been duplicated for decades but maybe it is something to do with interference from cystein.

 

Dissolve 3.55 g Na2HPO4 in 400 mL H2O in 500 mL volumetric flask. Add 7.0 g 2Na.2H2O.EDTA and 3.05 g cysteine.HCl.H2O. Adjust to pH 6.0 ± 0.1 with 1M HCl and dilute to volume with H2O. Prepare fresh daily.”



#2 mdfenko

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Posted 06 March 2014 - 05:31 AM

this is just a guess (and probably wrong, but...), casein is highly phosphorylated and will bind metals. a high concentration of edta should keep metals away from the casein.

 

also, the original investigator may have been using poorly deionized (or not deionized) water.


Edited by mdfenko, 06 March 2014 - 05:33 AM.

talent does what it can
genius does what it must
i do what i get paid to do

#3 DRT

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Posted 06 March 2014 - 11:04 AM

Ahh yes. Thanks.

I think you are correct; it would compete for the Ca and stop the casein forming micelles.






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