Easy purification of His-tagged extracellular secreted protein

Purification of proteins from eukaryotic cell supernatants is a tedious task. Normally, the concentration of extracellular secreted proteins is very low, and the proteins are easily degraded by proteases in the cell supernatant.

Of course, Cube Biotech Labs faces the same problem. One day, however, Cube Biotech scientists thought if we could add magnetic beads directly to the solution and isolate the His-tagged protein from the equilibrium system. Through the experiment, the entire purification process became easier than before. Therefore, we have developed a new method to purify His-tagged proteins secreted extracellularly.

Of course, the new method is successful because all of Cube Biotech's affinity purification beads: NTA, IDA, and the new INDIGO series can be processed in a high temperature steam sterilizer. Interested in trying our new method for the next expression of His-tagged extracellular secreted protein? Please refer to our operation flow below.

1. Take a vial of PureCube His label affinity magnetic beads (Ni-INDIGO, Ni-NTA, Ni-IDA) and remove the protective solution and add the same volume of PBS and then sterilize with high temperature steam.

2. Load the cell culture medium into the culture flask and inoculate the cells.

3. Add 100 μl (25% content) of magnetic beads treated with high temperature steam to the culture flask to 50 ml of the culture solution. The amount of magnetic beads can also be calculated by the amount of protein expressed.

4. After attaching a magnet to the outside of the culture flask, the cells are cultured according to the usual method. When the His-tagged protein is secreted into the supernatant, it will be captured by the magnetic beads.

5. When collecting protein, remove the culture solution and wash the magnetic beads fixed on the wall of the culture bottle with a cleaning solution.

6. Remove the magnet and transfer the beads to a clean tube.

7. Add the eluent and transfer the eluate to another clean vial.

8. The eluate contains the purified His-tagged target protein.

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