Figure 8. Peak overlays of all blue dextran elutions. Variance in peak height is due to variations in blue dextran spiking concentration and is irrelevant to the results.
As can be seen from the results, breakthrough volume was independent of flow rate, with a standard deviation of only 0.004 column volumes. This can be explained by the Van Deemter Equation:
\begin{equation} HETP=A+\frac{B}{u}+(C_{s}+C_{m})\bullet u\nonumber \\ \end{equation}
Where A is the constant eddy diffusion parameter, B is the diffusion coefficient in the longitudinal direction, and Cs and Cm are the resistance to mass transfer in the stationary and mobile phases, respectively. Because there is no mass transfer of blue dextran from the liquid phase to the solid phase or vice versa, this portion of the equation can be ignored. The flow regime is also operated at a relatively large u, making the longitudinal diffusion part of the equation negligible as well. This eliminates any dependence of velocity on peak broadening. These results are significant because they demonstrate a consistent breakthrough profile regardless of flow rate, which may need to be adjusted throughout the course of a continuous process. Therefore, breakthrough time across the column can be easily calculated from the current flow rate, column volume, and void volume coefficient. This is not necessarily true of other methods of flow through virus inactivation that rely on Dean vortices for axial mixing.
In order to demonstrate that live virus behaves similar to blue dextran, the column was spiked with inactivated xMuLV virus while running at a residence time of 30 minutes. Three 6 mL fractions of eluate were collected corresponding to before, during, and after the expected breakthrough of virus. Results can be seen in Figure 9.