Cell-Free Biologics Manufacturing: How It Works and Where It Fits

For decades, making a biologic has meant working with living cells. You develop a cell line, bank it, grow it in a bioreactor and harvest what it produces. Cell-free manufacturing takes a different route: it keeps the molecular machinery that cells use to build proteins and nucleic acids, but leaves the living cell out. The reaction starts when a DNA template is added, and it can finish in hours rather than weeks.

This guide covers the three forms of cell-free biomanufacturing now in commercial use, where each one fits, and what quality and regulatory teams should think about before adopting one.

What Cell-Free Manufacturing Means

Cell-free systems separate growing cells from making product. Cells are grown, then processed into an extract that retains the machinery for transcription and translation. That extract is characterized, stored and later used as a reagent. Alternatively, the components can be purified individually and recombined into a defined system.

Extract-based systems, drawn from E. coli, wheat germ, insect or Chinese hamster ovary (CHO) cells, are the productive workhorses. They are also less defined: extracts carry activities that were not deliberately added, which can complicate downstream work. Reconstituted systems, often called PURE systems after the approach described by Shimizu and colleagues in 2001, are built from purified components. They are cleaner and suited to research work, but yields are lower and they lack the chaperones and modification enzymes complex proteins need (New England Biolabs).

Reported outputs span a wide range. A 2021 review in BioTechniques collected figures from the literature: around 1 g/L for an antibody in an E. coli system, and reaction durations of about two days against at least a week for comparable cell-based expression (Chiba et al., BioTechniques, 2021).

Where It Is Being Used Commercially

Antibody-Drug Conjugates

Sutro Biopharma is the clearest example. Its XpressCF platform grows an engineered E. coli strain, processes it into a stored extract, and then expresses protein from added DNA in under 24 hours, with the same extract serving many different sequences (Sutro Biopharma, Form 10-K, 2024). The XpressCF+ version places non-natural amino acids at chosen positions in an antibody, giving defined conjugation sites. Sutro states this allows drug-antibody ratios above eight, where conventional cysteine-based conjugation is capped.

In January 2025, Sutro and Boehringer Ingelheim BioXcellence announced they had run the platform at 4,500 L at Boehringer’s Vienna site, with all batches meeting quality criteria for clinical use (Sutro / Boehringer Ingelheim, 2025). Sutro’s lead folate receptor alpha ADC, luveltamab tazevibulin, was deprioritized after a March 2025 portfolio review, but FDA cleared an IND for its next-generation tissue factor ADC, STRO-004, with first patient dosing expected before the end of 2025 (Sutro Biopharma, Q3 2025 results).

Conjugate Vaccines

Vaxcyte exclusively licenses XpressCF from Sutro for vaccine work and has partnered with Lonza on commercial manufacturing, including a custom suite at Lonza’s Visp site in Switzerland for its 24-valent and 31-valent pneumococcal conjugate vaccine candidates (Vaxcyte, 2023). That is a clear signal cell-free expression is moving into commercial vaccine supply.

Cell-Free DNA and mRNA

The second branch is nucleic acids. Touchlight produces doggybone DNA (dbDNA), a linear, covalently closed vector made enzymatically rather than by bacterial fermentation. In January 2025 the company announced an MHRA manufacturer’s authorisation covering GMP production of investigational API and critical starting materials for advanced therapies at its West London site, with claimed capacity above 8 kg per year (Touchlight / MHRA, 2025). Earlier, in February 2023, it reported the first FDA clearance of a US IND using dbDNA as an in vitro transcription template for mRNA in a cell therapy product, following FDA acceptance of a Drug Master File for GMP-grade dbDNA (Touchlight, 2023).

Elegen launched a cell-free synthesis product for IVT templates in September 2025, offering linear templates up to 5.5 kb in as few as 10 business days, with the poly(A) tail encoded in the DNA (Elegen, 2025). Both approaches target the same pain point: conventional plasmid DNA is amplified by bacterial fermentation, then purified and linearized, and plasmid purification is complicated by viscosity, shear sensitivity and the similarity between supercoiled plasmid and its own degradation products (IPS, mRNA manufacturing review).

It is worth remembering that IVT mRNA is itself a cell-free process. The enzymatic transcription and capping steps that make mRNA vaccines involve no living cells. What is usually not cell-free is the DNA template, which is why enzymatic DNA has attracted so much attention.

“Cell-free manufacturing does not remove biology from the process. It moves the biology upstream, into a reagent you characterize once and use many times.”

Advantages Worth the Attention

• Speed. Expression runs in hours rather than days, and a single characterized extract can serve many sequences, so no new cell line is needed per product. Compare that with the timelines in conventional cell line development.
• Difficult proteins. Proteins that burden or are poorly tolerated by a host cell can be made in an open reaction, because there is no cell viability to protect.
• Non-natural amino acids. The open format allows modified building blocks to be supplied directly, enabling site-specific conjugation chemistry.
• Screening throughput. Many DNA sequences can be expressed in parallel, which shortens candidate selection.
• On-demand supply. Freeze-dried reaction components can be rehydrated at the point of use. A 2016 Cell paper from the Wyss Institute reported a portable device making a vaccine antigen at an output equivalent to 33 human doses (PharmTech, 2016), a model relevant to outbreak response and remote settings.

Limitations and Open Questions

• Reagent cost. The energy molecules that drive the reaction are the dominant cost. This is the main reason cell-free has not displaced fermentation for large-volume proteins.
• Reaction lifetime. Reactions stall as byproducts accumulate, because there is no cell to clear them.
• Glycosylation. Bacterial extracts do not glycosylate. Eukaryotic extracts can, but with lower yields and higher cost. For glycoproteins where glycan structure drives efficacy or half-life, cell-based production usually still wins.
• Scale precedent. The 4,500 L milestone is notable precisely because it is unusual. Most cell-free work sits well below commercial biologics scale.
• Analytical burden. Extracts are complex reagents. Lot-to-lot consistency, host protein and nucleic acid clearance, and template removal all need method development.

Regulatory and Quality Considerations

There is no separate cell-free regulatory pathway. Products follow the same GMP and ICH quality expectations as other biologics, and reviewers have accepted them: FDA has cleared INDs using cell-free DNA and cell-free expressed candidates, and a Drug Master File exists for GMP-grade enzymatic DNA. What changes is where the control strategy sits. A 2021 review notes that GMP production and compliance with ICH Q5, Q8, Q9 and Q10 remain the gate to approval (Chiba et al., 2021).

Market Outlook

The market for cell-free protein synthesis products and services is still small. One commercial estimate puts it at about US$217 million in 2025, growing to roughly US$309 million by 2030, a compound annual growth rate near 7.3%, with Thermo Fisher Scientific, Danaher, GenScript, Takara Bio and New England Biolabs named among the main suppliers (MarketsandMarkets). That figure mostly reflects reagents and kits rather than therapeutic manufacturing revenue, so read it as a measure of activity rather than clinical impact.

What This Means for Biologics Developers

Cell-free is not a replacement for fed-batch CHO manufacturing, and anyone presenting it that way is overselling it. It is a focused tool.It earns its place where the molecule is hard for a living cell to make, where defined chemical modification is central to the product design, or where speed and the absence of fermentation materially change the supply model, as with DNA templates for mRNA.

If you are evaluating a cell-free route, treat it as a platform decision rather than a process change. Map the control strategy early, agree with your CDMO on how extract lots will be characterized and released, and discuss the approach with regulators before locking in a comparability plan. Precedents exist, but there are few of them, so the burden of explanation sits with the sponsor.

Terraforme Biosciences helps biotech companies and CDMOs build control strategies and regulatory submissions for novel manufacturing platforms, including cell-free expression and synthetic DNA. Learn more about our regulatory guidance services.