Regular articleIntegrated in situ-purification of recombinant proteins from Bacillus megaterium cultivation using SPION in stirred tank reactors

- Integrated product removal using functionalized SPION particle can be performed.
- Automated product removal of Protein A and α-lysozyme D1.3scFv antibody fragment.
- Product separation with SPION using handheld magnets led to high product purity.
- Aligning the magnetic poles lead to reduction in time of separation of up to 42 %.
- SPION particle in cultivation media does not negatively influence bacterial growth.

This paper discusses the use of two different types of functionalized superparamagnetic iron oxide nanoparticles (SPION) for automated in situ product purification of recombinant products directly from growing cultivations of Bacillus megaterium. Protein A as well as α‐Lysozyme D1.3scFv antibody fragment, both fused to a histidine tag, were successfully separated and purified from stirred tank reactor cultivations at lab scale. The SPION were separated using handheld neodymium magnets. Particle separation from the cultivation medium was investigated by magnetic field line simulation and validating experiments showing that a rapid and complete particle separation is possible. After 10 min of particle incubation the automated purification of 500 mL of cultivation medium was performed. The complete process, including the incubation time, was concluded within 30 min. 55 % of the total amount of Protein A and 83% of the total α-Lysozyme D1.3scFv amount were separated. Purity of both products was determined to be > 97.5 %. Very low particle concentrations of 0.6 mg mL−1 were sufficient for the separation. Bacterial growth was not influenced by SPION addition. This fast and integrated product removal for two different products shows the possible impact on automated large scale purification combined with the recycling of biomass.

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