Application-Driven, Top-Down Market
As a mature market, filtration moves forward incrementally through improvements in filter media and housings. “Developments are more on the application side than on the product side,” notes Karen Storm, director of sales and marketing at Sartorius Stedim Biotech (Göttingen, Germany).
Improvements in lab instrumentation and techniques demand a new level of sample preparation that was unnecessary a decade ago. High-performance liquid chromatography (HPLC) in particular has undergone a renaissance due to the advent of sub-two-micron columns. In an effort to reduce dead volumes while increasing sensitivity, system designers have eliminated all unnecessary volumes, thus placing a very high burden on sample purity. Particles that would simply flow through a 10-micrometer particle stationary phase get trapped within UHPLC columns or cause distortions inside detector cells.
“Since columns are so much more expensive, filtration and/or prefiltration of samples is mandatory in order to maintain column life in HPLC,” says Ms. Storm of Sartorius, “and for all analytical technologies as well.”
Throw it away
The last great technologic wave in filtration was the introduction of single-use or disposable products. Single-use filtration membranes and cartridges have become standard fare in large-scale processing in pharmaceuticals, biotechnology, food, and other regulated industries.
Disposables make sense for any process where cleaning and reuse must be validated. Apparent higher costs can often be justified on the basis of purified water and chemicals required to clean a multiuse filter and its housing, plus an operator’s time and the effort of validating the entire process. Eliminating cross-contamination is another obvious economic benefit, given the cost of a production campaign.
Since all these industries rely heavily on laboratory-scale analytical support, development, and scale-down modeling for troubleshooting, disposable filtration has caught on at very small scales. The idea is to reproduce exactly all manufacturing-scale operations, including filtration, at liter or even milliliter scale.
Disposables are significantly more expensive than multiple-use filters, either ceramic frit designs or membranes in stainless steel or plastic supports. “But pharmaceutical and biotech companies in particular are willing to pay premium prices to reduce hands-on time,” says Ms. Storm.
Considerations around cleaning, validation, and cross-contamination have trickled over to pure science and service labs as well, but a dichotomy still exists between academic and commercial labs. Universities, where labor is cheap, are more likely to reuse filters (even some models designed for single use), whereas companies generating results on which serious money depends increasingly prefer disposables. Every lab must do the accounting for its own workflows to be certain whether disposables or multiple-use filters make more sense.
Providing equivalent filtration equipment at large and small scale is now standard at large filtration companies. Similarly, process intensification— essentially eliminating steps—has become a buzzword both in manufacturing and during development. EMD Millipore (Bedford, MA) will soon launch a new line of depth filters that can eliminate centrifugation at the critical “clarification” step in cell cultures. The need for continuity between lab, pilot, and industrial scale is driving, at least in part, development of the new product line.
Clarification is the first step after cells are removed from the bioreactor. The process fluid is first treated with acid or a polymer flocculant to aggregate ruptured cells to cause them to clump together, thereby facilitating removal. “We realized that existing filters work better with cellular debris than with aggregated cells,” says Neil Soice, Ph.D., principal scientist at EMD. “These particles are larger than what you would normally see in filtration applications and would therefore rapidly plug the filters.”
The difficulty with centrifuges is that processes cannot be made fully disposable due to cleaning and associated validation. “Particularly at smaller scales, centrifuges are more trouble than they’re worth,” Dr. Soice adds. With the new depth filters, users need only flocculate the culture, filter, and concentrate before the first chromatography step, or sterile-filter the solution as is.
Tangential flow filtration (TFF), also called cross-flow filtration, is a relatively modern technique that involves causing fluid to pass across instead of directly through a filter membrane’s pores. The sweeping motion causes fluid to pass through and retains particles but significantly reduces clogging. TFF is used as an alternative to dialysis to concentrate large samples or process fluids containing proteins or viruses at the lab scale for biomolecules, nanoparticles, microspheres, or other particulates, and as a replacement for dialysis in buffer exchange.
But at small scale, centrifugation remains the method of choice for concentrating process fluids. Millipore’s Centricon® centrifuge filtration tubes are one such product. “The problem with centrifugation is that it requires a lot of fluid transfer steps, and it’s extremely time-consuming,” says Michael Bransby, western regional sales manager at Spectrum Labs (Rancho Dominguez, CA).
Spectrum has pioneered miniaturized TFF that concentrates up to 50 mL of particles or cells down to 500 µL in less than a minute. The technique involves a hollow-fiber cross-flow membrane in a tube fitted with several syringes. One syringe introduces the sample within the fiber, another collects fluid outside. A third syringe may be used to wash or collect the sample. In all, the process reduces the time to concentrate viruses or particles from a volume of 20 mL down to 500µL—a factor of 40—plus washing, from several hours to just a few minutes.
A group from the University of California, Los Angeles, published a paper late last year in the Journal of Virological Methods describing a technique based on TFF that concentrated a virus-containing sample by a factor of 2,000 in less than three hours.
Single-use cross-flow lab-scale filters are favorites in several industries outside pharmaceuticals and biotechnology, notes Karen Storm of Sartorius. The devices are used in water-purification and fish-breeding facilities to reduce volumes of water samples possibly containing viruses and to concentrate environmental samples as well.