A new scaleable freeze-thaw technology for bulk protein solutions.

A new method of freeze-thaw is described using experimental data obtained from freezing of purified rhGH (recombinant human growth hormone). The method is based on freezing protein solutions in rectangular rather than cylindrical containers. It is hypothesized that the change in container geometry allows for linear scale-up of the freeze-thaw operation based on equivalency of temperature-time profile. The hypothesis is tested using freeze-thaw data from a miniature (30 ml) and a 2.4 litre container. Computational fluid dynamics techniques are used to simulate the freeze process and the simulations are compared with experimental results. Protein quality is assessed as a function of freeze conditions using dynamic light scattering, circular CD, size-exclusion and reverse-phase HPLC measurements. The results demonstrate the applicability of the new approach. Freezing of rhGH solution at concentrations of approx. 30 mg/ml is shown to be possible with no damage to the molecule for up to five cycles of freeze-thaw. A nitrogen blast chest-freezer is designed and evaluated as part of the process. The refrigeration system and the freeze-thaw method can be used to freeze-thaw bulk protein solutions for development work and has the potential for transfer to manufacturing.