Ceramic hydroxyapatite requires special consideration during process-scale chromatography packing primarily due to its high specific gravity and rapid settling rate. Ceramic hydroxyapatite media have a bulk density of 0.63 g/ml and a free settling velocity of 35-125 cm/hr for 40 μm and 125-275 cm/hr for 80 μm. Additionally, ceramic hydroxyapatite media are sensitive to mechanical shear, which can fracture the particles and produce fines.
Bio-Rad InPlace columns offer a unique solution for packing media because they are designed with low-shear slurry valves, which minimize damage to media such as ceramic hydroxyapatite (Figure 1). Another feature of the Bio-Rad InPlace column slurry valves is their use for transferring media into and out of the column without removing the top flow adaptor. The valves are on the side of the column wall and do not interfere with flow dynamics.
The design of the Bio-Rad InPlace column allows a slurry-in-place operation so that the column can be repacked without removing the media or the top flow adaptor. With this procedure, the column can be repacked and ready for processing in less than 1 hr.
Unpacking of ceramic hydroxyapatite media can be challenging due to the high specific gravity of the particles, which form a dense packed bed. With a special unpacking protocol, ceramic hydroxyapatite can be unpacked from a Bio-Rad InPlace column in lass than 1 hr and using less than 2 CV of buffer.
In this study, six independent column packing cycles, including reslurrying, packing, and testing, were performed in less than 8 hr.