Hello Rashmi, How large are your protein monomer units? Are you expecting these units to have formed an oligomer? As a general rule of thumb, you want your protein molecules to be no smaller than 3x the membrane MWCO. Perhaps all you need is to try concentrating with a lower MWCO membrane, e.g. 3,000.
Alternatively, you could try: - change membrane manufacturers (this sometimes does make a difference), - pretreating your membrane (e.g. with BSA) to block all binding sites (but then you'll have to worry about BSA contamination of your sample), - concentrating your protein in under nitrogen pressure (I forget right now what the device is called but I used to use it all the time), - concentrating your protein in a speed vac to evaporate off some of the water, - ammonium sulphate precipitation, - or even TCA precipitation. HTH, C On Sat, Jan 28, 2012 at 10:54 AM, rashmi panigrahi < rashmi.panigrah...@gmail.com> wrote: > Hi all, > I tried to concentrate my protein using vivaspin 20 10,000 MWCO PES. > The protein was in 50mM Hepes pH 7.5, 500mM KCl and 10% glycerol. > I lost about 90% of my protein on the membrane of the centricon. > > Please suggest some way of concentrating this protein. > Will concentrating using peg 20K be a good alternative?? > > regards > > rashmi >
