We asked Dr. Tomohiro Nishizawa, who frequently purifies membrane proteins for crystallization, how he likes the new size exclusion chromotography column Superose 6 Increase. He is pleased with the shorter purification time and higher throughput compared with standard Superose 6 columns.
Purification time was the bottleneck
Using the Superose 6 column, I was able to perform fractionation on my membrane protein samples. However, low throughput was a major problem. For example, it usually takes one hour to purify one sample. Then there are hours of preparation and clean-up time. So it was difficult to purify multiple samples in a day.
We heard that the new Superdex 200 Increase has an extremely high resolution and an increased flow rate. So we gave the column a try to see if it would shorten the purification time. Membrane proteins tend to form micelles in detergents, resulting in a higher molecular weight. Unfortunately, the larger size pushed the target protein outside of the fractionation range for Superdex 200 Increase. The peak for the target protein came very close to the void volume peak area, so proper fractionation was not possible.
Now purification time is cut in half
Superose 6 Increase, on the other hand, has a fractionation range that is compatible with my membrane protein micelles. Therefore, the fractionation is going very well. I am especially satisfied because I can set a higher flow rate with this new column, resulting in a shorter purification time. I am surprised that I can now purify a protein in 30 minutes instead of an hour. This time savings allows me to purify three or four different samples in a day, which enables quicker mutant screening and faster results."
Method Purification system: ÄKTAexplorer 10S
Purification environment: 4°C
Sample: Membrane protein produced by insect cell expression system
Buffers: 10 mM HEPES, 150 mM NaCl, detergent
Learn more about Superose 6 Increase