DK <[email protected]> wrote: > In article <[email protected]>, Sudheer > Sangeetham <[email protected]> wrote: > >Hello all > > > >I would like to quantify the protein concentration. I checked the nanodrop > >manual in that they have given that I can estimate the protein by measuring > >directly at 280 nm by giving extinction coefficient value without doing > >bradford or lowry methods. I would like to choose measuring the protein > >concentration directly. Because I need to handle many samples all the time, > >so it is difficult for me to do bradford or lowry methods all the time. So > >How far is correct if I estimate the protein concentration directly? please > >give me your suggestion
> To expand on Nick's answer a bit: > 1. If your protein is quite pure, A280 + theoretical extinction coefficient > given by Protparam will *usually* get you as close to the real concentration > as almost anything else (quantitative amino acid analysis is still a golden > standatd but it's almost a lost art that few can execute competently and > reliably these days). > 2. If your protein is not very pure - particularly if contaminated by nucleic > acids or large amounts of small molecules that absorb UV strongly, then > just about anything else is going to be much more accurate than A280. To expand a little more: If the machine can reliably meassure at 205 nm that is the absorbtion maximum of the peptide bond. Takes care of the problem with a protein mixture. Small molecules and the buffer used might still pose a problem. -- Kaj _______________________________________________ Methods mailing list [email protected] http://www.bio.net/biomail/listinfo/methods
