Is it still possible to quantify cystein rich low molecular weight proteins such as Metallothionein in a given sample using Ellman's reagent if the sample is contaminated with some high molecular weight proteins?
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It's specific for thiols in general. If the contaminating proteins contain solvent accessible thiols, DTNB will react with them.
answered Jan 5, 2018 at 0:12
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$\begingroup$ Doesn't even need to be a protein. People in my lab often make large polymers with some thiol modifications. The thiols are quantified with Ellman's Reagent. $\endgroup$– user137Jan 5, 2018 at 7:23
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$\begingroup$ @user137 Yes certainly; I confined my answer to the question. $\endgroup$ Jan 5, 2018 at 7:31
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$\begingroup$ @canadianer How can we determine if the contaminating protein contains solvent accessible thiols? $\endgroup$– kash91Jan 5, 2018 at 22:39
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1$\begingroup$ @k.shekh First of all, it should be rather trivial to purify your low molecular weight protein from the contaminating high molecular weight proteins. I think that by far would be the best solution. If for whatever reason that’s not possible and you know the identity of the contaminating proteins, you can search the literature (including primary protein sequences) for information on the cysteine content. If the identity is unknown, you would need to make the determination yourself. I have had success labelling thiols with maleimide conjugated fluorophores and then visualizing them in a gel. $\endgroup$ Jan 5, 2018 at 22:50