Biomedical Engineering Reference
In-Depth Information
TABL E 7.4.
Impact of Column Diameter on Cost and Operation of the Protein A Column
Approximate
Resin Cost
(million dollars)
b
Number of
Chromatography
Cycles Required
c
Total
Processing
Time (h)
d
Column
Diameter (cm)
a
Resin Bed
Volume (L)
Eluate Pool
Volume (L)
140
308
3.1
13
10006
20
160
402
4.0
10
10053
15
200
628
6.3
6
9425
9
a
Twenty centimeters bed height.
b
Assumes an approximate cost of $10,000 per liter of rProtein resin.
c
Cell culture volume 15,000 L; titer 9 g/L; rPA resin binding capacity 36 g/L resin.
d
Linear flow rate 250 cm/h; processing time does not include support activities such as filter changes, buffer
preparation, and so on.
increasing concern, given that cell culture titers in excess of 5 g/L are becoming
increasingly regular [17]. At large scales and with the highly productive cell culture
processes, large columns are required to capture the entire quantity of product
produced in cell culture and rPA column can cost several millions of dollars.
Conversely, running multiple cycles on a smaller column increases the complexity
and duration of this step and, if taken to the extreme, can result in a significant product
throughput bottleneck.
To helpmitigate these concerns, flexibility in the size of the rPA columnwould allow
the cost and duration of this step to be tailored as the economics and throughput demands
of a product evolve throughout the product life cycle. For example, mAb2 has been
produced in cell culture at a titer of 9-10 g/L and exhibits a dynamic binding capacity on
the rPA column of approximately 36 g/L. The proposed design space for this product
would allow flexibility in the choice of column diameter and the number of chromatog-
raphy cycles run per production batch. The relationship between column diameter, the
number of chromatography cycles required, protein A resin cost, and processing time for
mAb2 is shown in Table 7.4. To ensure that these changes do not affect the process or
product, the critical and key parameters required to ensure consistent performance of the
column have been identified and will be maintained regardless of the column diameter.
For example, column-packing parameters, protein load, and mobile-phase composition
and linear velocitywould remainwithin acceptable limits regardless of column diameter.
In addition, the stability of the product in the capture column load and eluate is adequate
to allow for variation in the duration of this step associated with more or less column
cycling. Lastly, the volume (Table 7.4) and composition of the rPA eluate pool will not
significantly change, and therefore will not affect the performance of the subsequent
chromatography step.
7.6 HYDROPHOBIC INTERACTION CHROMATOGRAPHY
This third example pertains to purification of a FP, which employs a hydrophobic
interaction chromatography (HIC) step. Ammonium sulfate is added to the column load
