Not too long ago, cell-free technology—expressing proteins without the aid of living cells—may have seemed like science fiction. But in just a short time, cell-free expression systems have gone from a niche lab tool to industrial research and development pipelines. As its rapid growth continues, cell-free looks to become the new standard in synthetic biology.Cell-free technology is a powerful system for several reasons. With cell-free, proteins can be produced in a test tube—completely skipping the laborious process of using plasmids, lysing the cells, and purifying the mixture. The hassle of living cells (which are much pickier about their living conditions than test tubes and pipettes) is also avoided. Most critically, cell-free platforms lend themselves well to high-throughput screening, a necessity in the fast-paced, big data world of modern bioengineering.

The cell-free platform

Arbor Biosciences is a leader in cell-free space. When Dr. Evelyn Eggenstein (Division Manager, Synthetic Biology) and Matthew Hymes (Marketing Director) spoke with SynBioBeta earlier this year, the discussion focused on the technology’s availability and ease-of-use. When the two spoke with SynBioBeta again this month, it was clear Arbor Biosciences has quickly moved beyond proof-of-concept and is now working into the mainstream.Describing how the technology has grown, Eggenstein says, “We were able to work with customers on the industrial side who decided to implement cell-free technology through their R&D pipeline for screening purposes. screen a lot of constructs in parallel in a pretty short time. to go quickly from design to having the protein in hand.”To meet the growing demand for cell-free systems, Arbor Biosciences is adding bulk offerings of their master mix. Previously, the company offered 24 or 96 reaction kits for pilot studies, but that was not nearly enough for many customers. Now, 5-25 mL of their myTXTL master mixes are available for high-throughput applications, though larger orders are possible. These bulk options create the ability to screen thousands of constructs at the same time. But it’s not only the number of reactions or rapid screening that make cell-free platforms like Arbor Biosciences’ so exciting. Like any good biology system, a single component must work seamlessly with the whole.As the synthetic biology ecosystem grows, synergies between biotech companies become critical. One example is the Arzeda-backed platform collaboration between TeselaGen, Twist Bioscience, and Labcyte. Cell-free expression implemented in this integrated platform, accelerates the design-build-test cycle and data feedback loop. Optimization is essential as the speed of biotech innovation increases. “The convergence of all these next-generation technologies…, especially with AI and machine learning, has really accelerated the science,” says Hymes.

From proteins to building metabolic processes

As cell-free technology grows, so too do its possible applications, from simply producing a single protein up to the most complex scenario of generating bacteriophages.The possibilities, laid out in escalating order of complexity, start with protein or enzyme engineering-- where a single protein molecule is optimized to enhance its performance, and production yield or improve its biochemical characteristics. The next evolution in cell-free technology is to create antibody fragments, or other complex proteins which require a variety of chaperones and related elements for proper folding prior to use in downstream applications.These early knowledge gains give rise to establishing and designing full metabolic processes, through optimizing multiple enzymes and their interactions, in a single system. Metabolic process applications range from cannabinoids (requiring 3-6 enzymes) to biofuels (utilizing 8 or more enzymes).“Not only could you be using a cell-free platform to optimize each enzyme, you could also utilize cell-free expression to analyze an entire metabolic process,” says Hymes. This can lead to metabolically produced assays and ultimately, bacteriophages. “Knowing bacteriophages, with up to 300 genes in a genome, can be expressed in cell-free systems seems to imply there is no limit to how complex a final process can be generated in the system,” says Hymes.Cell-free systems are proving its worth; Hymes says the technology has moved from innovators to early adopters and is growing quickly. Of course, it’s impossible to know just when the technology will be fully mainstream, but Hymes and Eggenstein have high hopes for 2020. Both point to the fact cell-free technology has gone from having one session at a synthetic biology conference to having its own, dedicated, three-day conference. “That’s a big step in the technology development,” says Hymes.

Act locally, think globally

Where does this development eventually lead?The possibilities are seemingly endless. Industries across a range of sectors are looking for enzyme applications, from food and agriculture to soaps and cosmetics. Biofuels, of course, are another critical function as the world rushes to divest itself from hydrocarbons. There is also tremendous potential in diagnostics and therapeutics. Indeed, cell-free is already being used in paper-based forms like a pH strips to test water for viruses or to study the gut microbiome-host interaction for disease diagnostics.Eggenstein doesn’t just see the industrial potential—she believes cell-free can help shape the next generation of scientists. “It’s so much easier than using E.coli in the classroom,” says Eggenstein. Because cell-free systems aren’t alive, they don’t need special equipment, media, or any sort of cellular maturation phase. This enables synthetic biology experiments to be implemented into curricula of high schools and universities/colleges much more efficiently. “You really just need a simple pipette. Sometimes, you don’t even need an incubator—you can just do on the bench,” says Eggenstein.Synthetic biology looks to play a critical role in addressing our growing global crises, from food shortages to climate change. But as warming accelerates, our ability to research and develop new technologies must keep pace. Cell-free technology can increase the ease and speed of discovery and better prepare the next generation of scientists for the challenges ahead.