We'll be using TFF for two different tasks within the purification process: Concentration and Diafiltration. As the solution moves through the TFF apparatus, it leaves the supply tank...is pulled through a pump...past a pressure sensor...and then across the filter membrane.
Everything that passes through the membrane, including the buffer solution, is known as permeate - and for this process - is sent to waste. The GFP protein is larger than the pores of the membrane and is retained. The retained material - called Retentate - is recirculated to the supply tank. Recirculation of the feed continues until the desired concentration of GFP is achieved.
Following concentration, and while the protein solution is recirculating, a new storage buffer solution is added to the feed. In effect, the protein is being washed by the flow of a new buffer solution in, and the old buffer solution out.
As this diafiltration step proceeds, the buffer solution that’s being added to the feed replaces the buffer solution that the GFP was originally in, effectively removing any remaining salt as well.
When this process is complete, the GFP solution is routed through a point-two-two micron final filter...and then collected in appropriate containers - usually bottles or bags. The Purification process is complete. The Green Fluorescent Protein concentrate can now move downstream to final Fill-Finish to be freeze-dried and packaged.