Magnetic Bead Recovery Techniques

The photo sequence below demonstrates two ways, among many, in which a magnetic bioseparation can be accomplished.

Technique #1 (for small vessels):

  1. The magnetic agarose beads ( ~7 ml) having an attached ligand, are added to a graduated 50 ml plastic test tube and allowed to settle by gravity to measure their settled bead volume.

  2. A solution containing the target substance for the ligand (i.e. “ligate”) is added to the tube and the beads are shaken for several minutes - or whatever time is required for optimal binding between the ligate and ligand.

  3. A magnet is placed at the side of the test tube and in 1-2 seconds the magnetic beads will be seen to cluster- securely- along one side of the tube. The supernatant solution- now free of ligate- can be poured off.

  4. The tube is filled with a suitable wash solution or buffer and the tube is reshaken to resuspend the magnetic beads. After a minute, the magnet is again used to immobilize the beads at the side or bottom of the tube, allowing for the supernatant to be decanted -while the magnet is held firmly in place. A 2nd or 3rd such wash can quickly be done as needed.
  5. The tube is then filled with a solution which is designed to break the bond between the ligand and ligate, thereby freeing the latter into solution. The solution chosen is the same one that would be used in a comparable affinity separation which is done in a column.
  6. The magnet is placed at the side of the test , as described in steps 1.3 & 1.4, above, and the supernatant - CONTAINING THE RECOVERED LIGATE - is carefully collected and the magnetic beads are washed in water followed by storage (4-10°C) in a suitable storage solution such as 0.03 % sodium azide.

Technique # 2 (for large volumes of dilute ligate):

  1. A Neodymium Magnet (0.5”diameter x 0.25”thick) having a 5 lb. lift capability, is attached to a standard stir-bar retriever.

    Note: A typical stir-bar retriever does NOT have sufficient magnetic force to achieve good recovery of magnetic beads. After attachment of a small, Neodymium Magnet to its tip, however, it works nicely.

  2. A sufficient amount of ligand-attached magnetic agarose beads are added to the solution containing a dilute concentration of ligate. The volume of beads used is calculated based on the binding capacity of the beads in relation to the expected amount of ligate present in the solution to be harvested.
    1. The beads are gently stirred (overhead stirrer) or agitated ( roller bottle or equivalent).
    2. Note : Do Not use a magnetic stirrer or magnetic stir bar in an attempt to stir the magnetic beads - BECAUSE THEY WILL CLUMP AROUND THE STIRBAR AND MAGNETIC STIRRER AND NOT MIX PROPERLY.
  3. Sterilizing a sheet of Saran® or similar plastic film : In this procedure, the film acts as a removable magnet cover which allows the beads to be recovered quickly from a large volume of solution by moving the stir bar retriever/magnet assembly through the solution - thereby increasing the local magnetic field on the beads by bringing the magnet to them instead of expecting them to come to the magnet. This result is rapid recovery without the huge magnet that would otherwise be needed to attract all the magnetic beads contained in a large vessel.
    1. A sufficiently large sheet of Saran® Wrap or similar plastic film should be wrapped around the stir-bar retriever and Neodymium Magnet on it’s tip, such that it extends above the maximum point to which it will be immersed. The Saran® is then secured above the maximum point of immersion.
    2. The Saran®-wrapped stirbar retriever/magnet should then be placed in boiling water until sterilization is assurred. Alternate methods of sterilization can be used.
    3. The Saran®-coated stirbar retriever can then be immersed in the test tube or swept through a much larger volume and the magnetic beads are recovered in seconds.

      1. The magnetic agarose beads remain tightly bound to the Saran-coated stirbar retriever when the magnet is removed.

      2. The magnetically adhered beads can then be swirled in wash solution(s) followed by being swirled in a solution designed to break the bond ;between ligate and ligand -as with technique #1.
  4. The Saran-coated stirbar retriever is then placed in a sterile container and the film can then be peeled from the magnet such that the beads are either deposited in the sterile container or remain adhered to the outer Saran® surface - where they can be easily washed off with a gentle stream of water or buffer.

    Note: It requires virtually no force to peel the magnetic agarose beads away from the Neodymium Magnet at the tip of the stirbar retriever.

  5. The magnetic agarose beads can then be treated with a solution designed to break the bond between ligand and ligate - so the target substance can be recovered.

  6. The magnetic agarose beads can be recovered for future use by storing them in a suitable, refrigerated preservative (0.03% sodium azide).

Technique #3 :

  1. Simply place a suitable flat magnet underneath a flat-bottomed vessel containing the magnetic agarose bead suspension.
  2. In a short while, depending on the height of the vessel, the magnetic agarose beads will settle to the bottom- due to both the force of gravity and the additional magnetic force.

    Note: The larger the magnetic beads, the faster will be their rate of settling.