Thermo Scientific/Pierce has gotten into the protein purification game recently. The resin is slightly cheaper (USD $975/100 ml versus $1656 for GE) and the binding capacity is equivalent. I haven't rigorously run any regeneration tests though. Our local sales rep actually sent us a 1 ml spin column free of charge to give it a test drive.
As for the binding issues I've given up on flow through purification with GST. I lose too much protein. Nowadays I'll throw the lysate + extra protease inhibitor in a 50 ml tube with the resin and leave it on an extremely low speed rocking plate in the cold room overnight then load it into a column the next morning. The flow through is a cloudy mess and you have to wash it really well but all of the fusion protein is bound to the resin and my samples are stable after cleavage. Whatever works right? Cheers, Katherine On Wed, Jun 5, 2013 at 4:55 AM, Barbara Giabbai < [email protected]> wrote: > Hi all. > I don't have any experience with Clontech and Fisher resins, but about > the GE one I faced the same problem as Sebastiano indicated. > But I have to say that the problem was not general, but protein (or family > of protein) related: the low binding depends on the oligomerization state > of the chimeric protein. When it behaves as dimer (GST dimerizes) it's > fine, when the oligomeric state increases (in my case hexamer) I really > can't manage to purify the protein on GST sepharose. GST not exposed...I > guess... > > I'm waiting for the replies Mirek's question...I'm interested too. > > Ciao, > Barbara > > > > > > On Wed, 5 Jun 2013 09:32:28 +0200 > Sebastiano Pasqualato > <sebastiano.pasqualato@GMAIL.**COM<[email protected]>> > wrote: > >> >> Hi Mirek, hi all, >> I'm also very interested in the topic, so please keep me up with the >> replies, or make sure to post a summary, please. >> >> In addition to the price, the problem we're facing with GSH-beads from GE >> (although we haven't tried others yet) is that we can't manage to deplete >> our lysates. >> We are always left with a large amount of unbound GST-tagged protein in >> the flow through, that is eventually captured by a second, third and >> sometime fourth incubation with fresh beads. >> Using larger beads volume won't help. >> Has anybody faced and/or overcame this problem? >> >> Thanks a lot in advance, >> ciao, >> >> Sebastiano >> >> >> On Jun 4, 2013, at 8:54 PM, "Cygler, Miroslaw" <[email protected]> >> wrote: >> >> Hi, >>> I would like to ask the bb faithful for their experience with the >>> glutathione affinity resins. We have been using so far the Glutathione >>> Sepharose fast flow from GE but the price is getting steeper. We found >>> Glutathione Superflow resin from Clontech to be significantly less >>> expensive and Glutathione agarose from Fisher somewhere in-between. We have >>> no experience with the latter two resins and I wonder what is the >>> experience of other people with these resins? Do they have decent binding >>> capacity? Can they be efficiently regenerated or are they a single use only? >>> Thanks for your help, >>> >>> Mirek >>> >>> >>> >>> >> -- >> Sebastiano Pasqualato, PhD >> Crystallography Unit >> Department of Experimental Oncology >> European Institute of Oncology >> IFOM-IEO Campus >> via Adamello, 16 >> 20139 - Milano >> Italy >> >> tel +39 02 9437 5167 >> fax +39 02 9437 5990 >> >> >> >> >> >> >> > Barbara Giabbai > > Elettra-Sincrotrone Trieste S.C.p.A. > SS 14 - km 163,5 AREA Science Park > 34149 Basovizza, Trieste ITALY > Office: +39 040 375 8840 > Lab: +39 040 375 8537 > -- "Nil illegitimo carborundum"* - *Didactylos
