Early Pathogen Removal
Bioprocessors must beware of pathogens inherent in their expression systems, potential pathogens in ingredients and buffers, and those that enter the process from the environment or through worker contact.
Although viral clearance formally begins with the first purification step, definitive virus filtration occurs late downstream. Recent concerns regarding new viral and bacterial contaminants, however, have caused a whole-process rethinking of pathogen removal. For example Mycoplasma, the smallest known free-living and self-duplicating microorganisms, are particularly destructive and difficult to eradicate.
Genentech’s 2012 presentation on Leptospira contamination opened the eyes of many to the dangers of complacency with regard to novel organisms that have not previously been encountered in cell culture and fermentation. A poster, presented by Genentech scientist Ting-Kuo Huang, Ph.D., described the pathogen as “adventitious,” having been discovered “during routine microscopic visual examination.” Genentech considered the problem serious enough to warrant publicly exposing what is typically considered a trade secret.
“These concerns have led biomanufacturers to consider pathogen removal already at early process stages, which includes virus filtration, for certain cell culture media or media components,” says Holger Bromm, director of filtration technologies at Sartorius Stedim Biotech.
But as Bromm notes, removal of small contaminants requires finer retention characteristics of the filter media, which causes throughput to fall and cost to rise: “Cost is a critical issue here. For the future, it will be critical for vendors to offer filtration options that capture and retain smaller organisms such as mycoplasma, viruses, and other small contaminants without compromising current production costs significantly.”
Although virus filtration occurs late in purification, bioprocessors are uneasy with the notion of potential contamination persisting from one purification step to subsequent operations. “If you can detect contamination in your bioreactor, it is unacceptable to proceed further with downstream processing,” Bromm explains.
Chemical or heat treatments are two alternatives to filtration for eliminating early-stage contamination. But not all products and processes stand up to such treatment, and at their best they complicate the process by adding steps.