Automation is not suited to bulk preps for crystallographic work.
remillard, Post Doctoral Fellow, North America

Better antibodies against 6xHis tag.
jameslu, Principal Investigator, North America

Better solubility and yield of recombinant protein expressed and protease cleavage optimization.
amar, Graduate Student/Research Assistant, Asia

Better systems for expression of very large proteins.
amybio, Graduate Student/Research Assistant, North America

Competitive pricing, flexible and diverse multiple cloning sites.
cancun1991, Principal Investigator, North America

Cutting off the tag w/ TEV protease did not work at all for us. The enzyme degraded our protein (even though it does not have TEV sites except for between the tag and the protein) and never cut off the tag.
katiew, Principal Investigator, North America

Depends on the protein.
Mella, Principal Investigator, Central/South America

Development of more low cost methods.
jterner, Principal Investigator, North America

Expression yields remain the bottleneck, requiring massive scale-up in order to generate enough protein. SOLUBLE EXPRESSION remains the single greatest difficulty in drug discovery.
tstout, Lab Director/Supervisor/Coordinator, North America

Flow rate is unquestionably the greatest rate limiting step in our purifications.
cdominy, Lab Director/Supervisor/Coordinator, North America

I am using Amersham because that is what the authors of previous publications were using.
etoainshrdlu, Graduate Student/Research Assistant, North America

I am using Invitrogen's methanol-inducible Pichia pastoris expression system for recombinant protein secretion, hence don't use cell lysis procedures.
DeanM, Post Doctoral Fellow, Australasia/Pacific

I find that the new generation resins for chromatographic separation are extremely efficient and give very high yield of purified protein.
m__shyamal, Principal Investigator, Asia

I think the key event in the protein purification is the elution of the protein from the columns. It's at this stage when I find the yield varies between column makers, and also where the companies should focus to formulate more efficient eluting buffers.
chivite, Graduate Student/Research Assistant, Europe

I would appreciate more protocol links on protein purification.
Sivvy34, Graduate Student/Research Assistant, Asia

I would like to have more facilities to get protein expression after cloning my gene in an expression vector. Sometimes, but not every time, I do clone a gene in an expression vector but I do not obtain any expression, without any reason.
netorino, Principal Investigator, Central/South America

Improved non-specific adsorption.
Nice, Principal Investigator, Australasia/Pacific

Invitrogen should reduce their prices.
T.Rat, Post Doctoral Fellow, Europe

Is there a workstation that can automate gel filtration, ion exchange and affinity chromatography?
Venk, Professor/Teacher, North America

Lower cost of chromatography supplies
metacat, Post Doctoral Fellow, North America

Lower prices for the reagents we use
cruiser, Professor/Teacher, North America

Make it easy to routinely get purifications of protein to >95% purity in one step.
wtaylor, Lab Director/Supervisor/Coordinator, North America

Not everyone is going to have their own FPLCs. So a small individual FPLC-like piece of equipment around $5-10K would sell really well both in academic and pharma. If you want a similar example, look at the Guava personal FACS machine. I know of six labs in my company that own one.
kemtoi, Staff Scientist, North America

Our preferred expression host is E. coli, because of ease and cost. The downside is that it's not a great host for many mammalian proteins, especially larger ones. A modified host better able to deal with > 80 kDa human proteins would be great!
Digard, Principal Investigator, Europe

Overall, I am satisfied with the products and support for our supplier, Qiagen. We have only recently run into problems in trying to tag a large protein, and can not say with certainty if a longer his tag or different tag would work better (I don't have the time, nor the resources to explore this further). We have occasionally run into problems with protein solubility using hig tagged proteins, which we solved by using NEB's MBP fusion vectors. We would be tempted to try the IMPACT system, but it's too pricey at the moment, and we may try it at a later date.
szat, Professor/Teacher, North America

Phage display is becoming huge. I think it would be great to see more kits available for quick panning, screening, and subsequent expression. Our in-house vector is clunky and laborious to use. But at least our expression is low (sarcasm intended).
lahn1, Staff Scientist, North America

Proteins are isolated from 1 liter of cleared insect media, limiting the number of preps to 1 at at time. My proteins of interest are quite large (>200 kDA), and the expression system we have been using exports at least 2 other proteins of a similar size.
dfritzin, Principal Investigator, North America

Purification of recombinant proteins is focused on speed of purification not on purity. Purity is often only tested with SDS-PAGE and CBB staining often in suboptimal arrangement. Especially in automated systems purity has to be improved.
Wojtek, Lab Director/Supervisor/Coordinator, Australasia/Pacific

So far all the 6xhis antibodies I have tried are very poor. Companies should offer the same vector with different promoters. Some promoters are more tightly controlled than others.
Proteome, Staff Scientist, North America

Solubility tags, bacterial secretion systems, alternative bacterial expressions systems that are already well developed (Bacillus subtilis, Aspergillus niger).
cchang2, Post Doctoral Fellow, North America

Suppliers simply have to have a closer look to the real needs of pharmaceutical companies and not to offer a package that has been designed to be on the market ASAP. As a matter of fact, we industrials have to adapt too much to their purification systems, instead of the opposite.
nony, Lab Director/Supervisor/Coordinator, Europe

Thank you very much.
sraju, Principal Investigator, North America

The ability to produce a purified protein product is an arduous task. Problems stemming from cloning, to expression, and to final yield and activity are always stopping steps in the experiments completion. One problem I have always had with protein purification is the lack of functionality. I wish there were better buffers and prep systems made to increase protein biological activity.
Gruther, Staff Scientist, North America

The biggest problems arising from E coli hosts are (a) proteolytic activity (b) solubility of expressed protein (in our experience Maltose binding protein gives the best results) and (c) the price of scaling up affinity chromatography for protein purification. Proteolytic activity is the biggest problem since frequently we observe impurities resulting from fragmentation of the protein of interest.
jacksc, Staff Scientist, Australasia/Pacific

The most important thing in purification is purity, however, there always has some unspecific cleavage during the tag removal process, it affects the purity and the production greatly. If you could include cleavage in the automated protein purification system, it would be big progress. Of course, it should appear as an option.
y27xiang, Post Doctoral Fellow, North America

The NTA resin is the one we use right now. We have already tried every other resin on the market and this one works the best with the protein that we purify. Although it works, it does not bind all of the available protein and there is room for improvement.
eharri, Post Doctoral Fellow, North America

The protein purification that I conduct in on a large scale (1kg per run).
gordorito23, Production/Manufacturing, North America

We are considering an automated system like €kta, but we haven't decided yet.
tblumens, Post Doctoral Fellow, North America

We need a 'screening' automated system that will produce ~ 1 mg protein for initial studies, but we'd then want to scale up selected hits to produce ~ 50mg per run.
rgrant, Post Doctoral Fellow, Europe

We perform old-fashioned "bucket biochemistry" to purify various types of collagen from fetal bovine tissues. These methods require volumes of buffers on the order of hundreds of liters at a time.
dbrand, Principal Investigator, North America

We use Precission Protease from Amersham. Higher flow rate is always a plus. Robust column, which can be run at high speeds on HPLC and FPLC (i.e., 10 mls/min). Columns, which are highly resistant to harsh chemicals --facilitating column cleaning.
chong, Graduate Student/Research Assistant, North America

We use Prescission Protease from Amersham.
dmwalter, Staff Scientist, North America

We use the MBP-tagged expression system equally as often as the GST- tagged system. We have found this an equally useful method for preparing our fusion proteins.
kellerizer, Professor/Teacher, Australasia/Pacific