Which buffer is used when transferring into the microcentrifuge tube with supernatant?

The binding buffer is crucial in the process of purifying proteins such as Green Fluorescent Protein (GFP). During the purification process, once the cells have been lysed and debris has been removed, the supernatant contains the soluble proteins. The purpose of the binding buffer is to promote the binding of the target protein—in this case, GFP—to the chromatography resin or affinity column.

The composition of the binding buffer is specifically designed to optimize interaction between GFP and the resin, enhancing the efficiency of the purification process. This involves maintaining a suitable pH, ionic strength, and possibly including components that stabilize the protein structure, ensuring that GFP retains its functionality and affinity during the purification steps.

In contrast, the other buffer types serve different roles. The equilibration buffer prepares the resin for binding, the wash buffer is used to remove non-specifically bound proteins after the initial binding, and the elution buffer is employed later to detach the target protein from the resin for collection. Each buffer has a distinct function that is critical to the overall protein purification workflow, but the binding buffer specifically is what facilitates the initial capture of GFP in the microcentrifuge tube with the supernatant.