Tangential flow filtration (TFF) is a useful method for separating molecules or particles in liquid solution by size and enables both concentration and the ability to change buffer composition.1 TFF is used in almost every biopharmaceutical manufacturing downstream process to provide gentle, fast concentration and diafiltration—a dilution process where components are removed or separated. In comparison, gel filtration chromatography separates molecules by size and provides the ability to change buffer composition, but this technique cannot concentrate the target entity.
In normal flow filtration (NFF) solids often form a cake or a second layer over the membrane. When processing biomolecules or smaller particles even a thin second layer on the membrane is often enough to foul it, reducing or stopping the filtration process.
Optimizing parameters
“Pressure and flow are the key parameters to control and monitor,” says Fredrik Lundström, senior product manager at Cytiva. “It is strongly recommended that pressure be monitored on all three streams: feed, retentate, and permeate.”
The filter membrane area should be utilized as efficiently as possible, and the process optimized to operate at a high flux (permeate flow/membrane area) that does not foul the membrane. “There is a balance of applying sufficient transmembrane pressure (TMP) to achieve a high flux, but not so high that the membrane fouls,” says Lundström.
To prevent fouling for microfiltration operations, where the filter pores are large, controlling and limiting the permeate flow (flux) is key. In ultrafiltration, where the pores are small, it is important that the TMP is not too high.
According to Lundström, if the process includes diafiltration and buffer exchange, it is important to optimize the diafiltration starting point—the amount of target product to be concentrated—to minimize buffer use and filtration time. A continuously fed diafiltration buffer to the TFF recirculation container is more efficient and saves time and buffer.
“It is important to monitor the buffer weight and product containers to determine the concentration factor—how many times the product has been concentrated—and the diafiltration factor—the number of diafiltration buffer exchanges that the product has been exposed to,” recommends Lundström.
Different approaches
TFF is a classical bioprocessing operation for harvesting, clarification, and final concentration with buffer exchange. TFF is also used for novel modalities like viral vectors or mRNA applications. In these applications, the filtration technique is more prevalent early in the process than for traditional mAb processing.
There are two common types of TFF devices: cassettes and hollow fibers. Cassettes are commonly applied for mAb concentration as they provide a higher flux, and hollow fibers are useful when the target entity is shear-sensitive. “This has made hollow fibers more interesting with the increased need for processing cells and shear-sensitive viruses like lentivirus,” said Lundström.
The TFF technique can also be implemented as continuous cultivation (perfusion), either in production or an N-1 bioreactor, where it can provide additional controls compared to alternating tangential flow filtration (ATF).
When TFF filter cassettes are placed in series it allows concentration with a single pass of liquid over the TFF membrane instead of multiple times. “In this instance, since single-pass TFF does not require a batch type of operation, it has generated a lot of interest in the continuous processing area,” says Lundström.
Dual cyclic TFF systems2 where a peristaltic pump feeds the stream through two membranes with pore sizes of 200 and 30 nm is deemed a better option for isolating extracellular vesicles in the 30 to 200 nm size range, compared to direct filtration or single cyclic TFF. Combining buffer exchange with dual cycle TFF further aids in eliminating contaminants.
Advice for TFF applications
“First consider the type of application and what the target of the unit operation is such as concentration factor, buffer exchange, target entity, and liquid composition. This can give you an indication of which type of TFF device and the pore size of the membrane you should start with,” says Lundström. “Then you can start screening different filters.”
Filter suppliers often have process parameter recommendations such as cross flow for a particular filter type to use to run initial tests. “You should vary the TMP and measure flux to find the optimal processing conditions,” advises Lundström. “Other parameters, such as the temperature of processing, may also need to be considered. It is also beneficial to use smaller lab and pilot scale systems to screen filters and optimize the process.”
TFF solution suppliers are available to help with the filtration process, membrane selection, and provide guidelines for processing conditions. If the process will be scaled up the availability of filters and systems for larger sizes should be considered.
References
- Musumeci T, Leonardi A, Bonaccorso A, Pignatello R, Puglisi G. Tangential Flow Filtration Technique: An Overview on Nanomedicine Applications. Pharm Nanotechnol. 2018;6(1):48-60. doi: 10.2174/2211738506666180306160921. PMID: 29510657.
- Kim K, Park J, Jung JH, Lee R, Park JH, Yuk JM, Hwang H, Yeon JH. Cyclic tangential flow filtration system for isolation of extracellular vesicles. APL Bioeng. 2021 Feb 10;5(1):016103. doi: 10.1063/5.0037768. PMID: 33688618; PMCID: PMC7932758.
