The channels of a monolithic column allow convective mass transfer between the mobile phase and the resin. This property of monoliths simplifies the scale-up process as retention time does not affect the resolution and dynamic binding capacity of the column. Consequently, the bed height of CIM® columns can be kept very short.

The short bed height minimises the backpressure allowing high flow rates even at production scale. Existing skids can achieve of 30–60 CV/h on a CIM® 8 L column. In current gene therapy manufacturing, 8 L column is the largest unit used in production of viral vectors, plasmids, and other biomolecules for clinical applications.

Iso-resolution scale up on CIMmultus™ preparative columns

Linear gradients are generally used in purification of large biomolecules. Milavec et al derived a very simple equation for scaling up/down an optimized gradient, maintaining the resolution during the process. For starting column 1 and target column 2:

Gradient time

Where tg is the gradient time, V is the column volume, F is the flow rate and L is the bed height, calculated as half the difference between the outer and inner radii of the column (found in Table 1).

This approach allows the flexibility of varying the flow rate during scale-up and is enabled by diffusion-free mass transfer in monoliths. It is important to note that some fine tuning of the gradient may be required after the method is transferred.

table 3
Table 1: Information about CIMmultus™ columns

In the following example, a mixture of three proteins (myoglobin, conalbumin and trypsin inhibitor) is separated in a linear salt gradient on an 8 mL monolith column. The method is scaled up to 80 mL and 800 mL columns using the equation above.

Figure 4: Separating a mixture of proteins in a linear salt gradient, scaled up from 8 mL to 800 mL. The resolutions between peaks 1 and 2 are 2.98, 3.0 and 2.98 for each volume in sequence, and the resolution between peaks 2 and 3 are 3.14, 3.1 and 3.23 for each volume in sequence.

BIA Separations columns are pre-packed in selected volumes. Bed height, or thickness of the monolith tube in the case of CIMmultus™ preparative columns are fixed values and do not need to be considered when scaling up. The methods presented above are applicable to most scenarios, even when delicate separations are required.


  • Žmak, P. Milavec, et al. "Transfer of gradient chromatographic methods for protein separation to Convective Interaction Media monolithic columns." Journal of Chromatography A 1006.1-2 (2003): 195-205.