Dealing with Leachables
Gerhard Greller, Ph.D., R&D director for upstream technology at Sartorius Stedim Biotech, focused on the optimization of single-use bioreactors. With the widespread adoption of single-use systems, the problem of leachables from the materials used to construct the plastic is becoming more significant.
“If you are ordering 50 L of media at a time, you may need to store it for weeks,” explained Dr. Greller. “That means these substances can drastically affect the growth of cells.”
Dr. Greller and his team have identified many of these compounds and redesigned the film that lines the bag. “We had to start from the resin selection. Through this approach, we went through a rapid learning curve, allowing us to characterize the leachables, including materials released by the gamma-irradiation process.”
Dr. Greller mentioned the work of Matthew Hammond and his coworkers at Amgen, who have identified one of the most toxic extractable compounds from a number of substances present in the polymer films used in presterilized, single-use biomanufacturing systems.
This molecule, bDtBPP, is detrimental to the growth of a number of strains of CHO cell lines, even at concentrations as low as 100 µg/L. It is derived from the compound tris(2,4-di-tert-butylphenyl)phosphate, and affects the mitochondrial membrane potential. Ionizing radiation, used in the sterilization process, appears to be the mechanism by which bDtBPP is generated.
Another critical quality control issue emphasized by Dr. Greller is the monitoring of leakage through suitable testing programs. The test method must effectively identify potential damage to installed bioreactor bags, including the bag seals, port welds, connections, and bag surfaces, which could be damaged during their assembly. These failures could cause substantial financial losses as well as constituting a significant safety hazard.
To deal expeditiously with the need for effective leak monitoring, Dr. Geller’s colleague, Martin Dahlberg, uses the Sartocheck®4 Plus Bag tester with a bag-tester fleece that prevents masking of any leaks that may have been introduced during installation.
“It allows point-of-use leak testing of single-use bioreactor bags post-installation and pre-use in its final bag holder,” said Dr. Geller.
The goal of these advances is to ensure the end user does not have to touch the bag and accessory components any more than necessary.”