Impact of Lot-to-Lot Variability of Cation Exchange Chromatography Resin on Process Performance - A case study to understand the impact of lot-to-lot variability of a cation exchange resin on process


Impact of Lot-to-Lot Variability of Cation Exchange Chromatography Resin on Process Performance
A case study to understand the impact of lot-to-lot variability of a cation exchange resin on process performance.

BioPharm International
Volume 21, Issue 5

Column Evaluation with NaCl

Table 1. Column asymmetries
Table 1 shows that the column asymmetries, evaluated with 1 M NaCl, varied from 1.0 to 1.7.4 The table also points to a correlation between column asymmetry, as evaluated with NaCl and elution volume. Generally, as asymmetry increases, column volume also increases. This suggests that the columns may have been poorly packed, resulting in varying flow patterns. To determine if this was the case, the probable correlation was investigated using two methods.

Figure 5. Peaks from acetone injections
Column evaluation with acetone. In the first method, ionic interactions between the analyte and the resin were eliminated by using a 3% v/v acetone injection. Acetone should not interact ionically with the cation exchange resin. Column asymmetries when evaluated with acetone were almost identical (Figure 5). Acetone evaluation showed that in the absence of ionic interactions, all four columns showed equivalent flow characteristics.

Figure 6. Elution with high conductivity buffer
Elution with high conductivity buffer. The second method of investigation used a high conductivity buffer to elute the bound antibody off the columns. Columns containing cation exchange resin lots A and D were used because they represented the extremes of asymmetry and column volume. The purpose of this experiment was to see if the different shapes of the elution peaks were caused by poor column packing or if they were because of different selectivity of the resin for the charged isoforms of the antibody. The use of a much stronger elution buffer would significantly reduce the selectivity for all the isoforms so that if column packing was predominantly responsible for the different peak shapes, these differences would still be observed.5 The chromatograms are shown in Figure 6.

The solid lines show the widely varying elution profiles and volumes that were produced when the normal elution buffer was used. The dashed lines show that with the high conductivity buffer, the elution peaks were almost identical. The fact that no difference in the elution profile was observed under high conductivity conditions indicates that the variation in the elution under normal conditions is largely because of the different lots of resin having a difference in selectivity for the charged isoforms of the antibody. Thus, this variation is not likely to be related to column packing. To further understand the reason for this different selectivity, we examined the ionic capacity and the particle size distribution of the four resin lots.

Ionic Capacity

Table 2. No variation in ionic capacity
Certificates of analysis from the manufacturer contained the ionic capacity of each lot of resin. Table 2 lists the ionic capacities of each of the four lots. They are very similar and no trends are apparent. Ionic capacity of the resin lots did not cause the lot-to-lot variability in peak shape and elution volume that we had observed.

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