Development of a Novel Platform TFF System for Insect Cell Culture Harvest - Development of a Novel Platform TFF System for Insect Cell Culture Harvest A single-membrane system was used to clarify ins

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Development of a Novel Platform TFF System for Insect Cell Culture Harvest
Development of a Novel Platform TFF System for Insect Cell Culture Harvest A single-membrane system was used to clarify insect cells, diafilter the cell concentrates, mix with an elution buffer to release the target protein, and collect the released target protein in the filtrate. The one tank–one module method simplifies and improves the harvest in comparison to multiple centrifugation and filtration steps.


BioPharm International


Despite the lower flux rate, using only one module has several advantages over using one module each for clarification and protein extraction. First, buying only one module reduces capital expenses. Also, because the module does not need to be cleaned or changed between the clarification and protein extraction steps, the process time necessary to clean or replace the module is saved. In this case, only 30 min. were needed for the extraction buffer and cells to circulate. In contrast, the time to clean, install, and rinse a new module would be approximately 2 h. In addition, in a commercial process requiring good manufacturing practices, cleaning validation would only need to be performed once instead of being performed twice (once for each of two membranes). Over the lifetime of the process, this reduction in cleaning validation could eliminate the need to validate the module hundreds of times. Another advantage of this setup is that the cell solution is not removed from the tank. This reduces the chance of spillage or contamination.

Scale-Up

One advantage of using TFF over centrifugation is that TFF scales up linearly.6 Microfiltration TFF is scaled by maintaining a constant ratio of filtrate volume to membrane surface area. The channel height, membrane material, pore size, and shear rate are kept constant. Based on the results obtained in this study, a 1,000-L batch could be harvested and processed in the same manner as described here in 2.5 h using one hundred times the membrane surface area than the initial trial. If a higher protein recovery yield is desired, the operational time would increase to 4.5 h to recover 99% of the protein. This scale up could easily be achieved by using an Optisep 11000 holder with 20 m2 of membrane. A shorter process time could be achieved by increasing the membrane surface area.

Summary


Quick Recap
A new protocol for the clarification, extraction, and concentration of protein from insect cells has been presented. This procedure is a modified tangential flow filtration process, "SFTFF," which uses a single module for the clarification and extraction of the desired protein. Using one module for these two steps saves both time and material costs. Finally, diafiltration was performed to remove small proteins and medium components from the protein. Thus, the protein can be placed in a buffer that permits loading onto the chromatography column, which saves processing steps.

RICHARD CHUBET is a scientist at Protein Sciences Corporation, 1000 Research Parkway, Meriden, CT 06450, 203.686.0800, fax 203.686.0268,
JAMES KACMAR, PH.D, is a process development scientist, and HANK KOPF is chief technology officer, both at NCRST, Inc., 1000 Goodworth Dr., Apex, NC 27539, tel. 919.387.8460, fax 919.387.8540,

References

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