The Äktapilot from GE Healthcare (formerly Amersham Biosciences) is an easy to use protein purification system, enabling protein production on a gram scale. It is FDA approved (for validation), completely sanitizable and purification can be performed under GLP and cGMP conditions. The UNICORN software that operates the system makes it very easy to control the Äktapilot. The provided “method wizard” allows one to logically develop a fully automated protocol. The same software is also used to control lab scale purification systems, such as ÄktaExplorer. As a result, once a small-scale purification method has been optimized, it can easily be transferred to the Äktapilot system for production scale purification.
During the purification process one can assess several parameters including pressure, conductivity, pH and UV absorbance (simultaneous measurement of three different wavelengths). This feature allows detection of a contaminant provided it has a different absorbance spectrum than the protein of interest. Furthermore, the system has two air sensors; one immediately before the column, preventing air bubbles from entering the column and the other for air bubble detection during sample application. The system has also has two (buffer) pumps for gradient generation and a third pump for sample application that allows the user to have a fully automated protocol. The buffer pumps can also be used for sample application to the column, offering two advantages. First, when two pumps are used the maximum volumetric flow is twice as fast (i.e. instead of 24 l/h, one can increase the maximum to 48 l/h). This translates to a processing time of 100 liters in about 2 hours. Based on my experience, I would recommend using only 75 % of the maximum speed, which allows you to completely process 100 l of sample in less than 3 hours (leaving enough time for washing, eluting and cleaning within one day). The second reason for using the buffer pumps is that the air trap filter can be placed in-line, preventing air bubbles formed during cold storage from getting into the system.
After sample application and washing of the column, fractions can be collected during elution in several ways: 1) automatically, using the 8 outlet valves provided, 2) manually, or 3) using a fraction collector (optional). When I use the fraction collector (Frac950; Unicorn software controlled) it is typically for small-scale testing using a 20 ml column. This is an easy method and gives good reproducible results for scaling up. Changing the columns and calibrating the pH electrode is a bit inconvenient due to the need for UV fiber optics transfer, which is housed on the outside of the system. Overall, the Äktapilot is a very practical and easy to use protein purification system.
Arnold Vos
AVBIOTEC
Protein Engineering Consultant