Welcoming ExpiCHO-S to our mammalian expression portfolio

The Protein & Structure department at Sygnature Discovery are excited to announce ExpiCHO‑S^TM as the latest addition to our mammalian expression platform. This expansion of cell line capabilities further enhances the flexibility and scalability of the recombinant protein expression services we provide to clients across biotech and pharma.

As part of the onboarding process, we compared expression of various test proteins, including 4 antibodies in ExpiCHO‑S^TM with our well-established CHO-3E7 cell line and we present that data below.

Choosing the Right Cell Line Matters

It has always been our philosophy to choose the expression host cell based on the nature of protein target itself and where there are multiple possible options, to perform a comparison screen early in a programme. Different targets respond differently to each system, and selecting the right platform from the outset can:

• improve yields
• reduce development timelines
• minimise costs
• ensure delivery of the required protein quantity and quality

Within our Cell Science team, we offer a diverse suite of eukaryotic expression systems to support proteins of varying complexity and production needs. Our portfolio now includes:

• HEK293‑6E and CHO‑3E7 (both licensed from the National Research Council, Canada)
• Expi293™ and ExpiCHO‑S^TM (both licensed from Thermo Fisher Scientific)

Why do we need mammalian expression platforms?

Mammalian expression platforms are essential for producing proteins that closely replicate their natural human counterparts, making them vital for both research and biotherapeutic development. Unlike bacterial or insect systems, mammalian cells provide the full repertoire of human‑like post‑translational modifications (PTMs)—particularly glycosylation, disulfide bond formation, and complex folding pathways—that many secreted, membrane, and multi‑subunit proteins require to be stable and functional. These systems also offer the appropriate lipid environment, chaperones, and trafficking machinery needed for challenging targets such as GPCRs, cytokines, and antibodies. As a result, mammalian platforms such as CHO and HEK293 have become the industry standard for generating biologically relevant and regulatory‑compliant protein material, underpinning the production of most approved therapeutics and enabling high‑fidelity studies of protein structure and function.

Comparing ExpiCHO-S^TM to CHO-3E7

With an increasing demand for high‑yield mammalian expression systems, we evaluated ExpiCHO‑S^TM to understand where it fits relative to our existing mammalian platforms.

For this comparison we selected a range of test proteins

• TIMP2 is a well‑characterised secreted protein, which we have produced extensively across several cell lines. TIMP2 exhibits intermediate expression levels, making it an excellent benchmark for detecting improvements or shifts in productivity between systems.
• Two mCherry tagged proteins, one intracellular and the other that is secreted
• Four recombinant antibodies: Depemokimab, Bococizumab, Eldelumab, and Sacituzumab

For each of these proteins we compared:

• CHO‑3E7, as a control using our internally optimised expression workflow
• ExpiCHO‑S^TM, using the Thermo Fisher ExpiCHO™ Expression System Kit

Overview of the ExpiCHO-S^TM protocols

Alongside the cells Thermo Fisher also provides three recommended expression protocols for ExpiCHO‑S^TM—called STANDARD, HIGH, and MAX—each incorporating different feeding and temperature‑shift strategies to enhance protein yield.

STANDARD protocol:

• Transfection at 37°C
• Addition of enhancer and feed at 24 h post‑transfection
• Culture maintained at 37°C to harvest (Day 8-10)

HIGH protocol:

• Same transfection/enhancer/feed schedule as STANDARD protocol
• Temperature shift to 32°C after the 24 h feed
• Culture maintained at 32°C to harvest (Day 10-12)

MAX protocol:

• Same transfection/enhancer/feed schedule as STANDARD/HIGH protocol
• Temperature shift to 32°C after the 24 h feed
• Second feed (day 5)
• Culture maintained at 32°C to harvest (Day 12-14)
• Designed to maximise productivity

TIMP2: Greater yields observed across all ExpiCHO-S^TM protocols

Our evaluation of secreted TIMP2 demonstrated a marked improvement in expression across all three supplied ExpiCHO‑S^TM protocols (Figure 2, left). Each expression workflow—STANDARD, HIGH and MAX—produced substantially higher titres than our established CHO‑3E7 process for this particular protein, highlighting the robustness of the ExpiCHO‑S^TM system even without any additional optimisation.

mCherry secreted construct

The trend seen with TIMP2 was further supported by our secreted mCherry control construct (Figure 3, right). The parallel increase in expression for both TIMP2 and mCherry indicates that the observed improvements are system‑driven rather than target‑specific.

Together with the TIMP2 data, these results strongly suggest that ExpiCHO‑S^TM, used in combination with the supplied manufacturer‑recommended protocols, represents an excellent addition to our mammalian expression portfolio for secreted proteins and provides clients with a high‑yield alternative to our existing CHO platform.

mCherry intracellular construct

Interestingly, the expression of an intracellular mCherry control construct did not show a comparable improvement in ExpiCHO‑S^TM cells (Figure 3, left). When benchmarked against our in‑house CHO‑3E7 platform, intracellular mCherry levels remained broadly similar, suggesting no clear advantage to switching systems for cytosolic targets. Given the higher media and transfection reagent costs associated with ExpiCHO‑S^TM, these findings indicate that CHO‑3E7 may remain the more cost‑effective choice for intracellular protein production.

Recombinant Antibody Production: Strong Performance in ExpiCHO‑S^TM

Finally, we evaluated recombinant antibody expression in ExpiCHO‑S^TM and found that this system can achieve yields approaching 1 g/L, underscoring its suitability for demanding biotherapeutic projects.

Using the Thermo MAX protocol (described above), we expressed His‑tagged Depemokimab and Eldelumab, followed by nickel‑affinity purification. Both antibodies showed substantially higher yields in ExpiCHO‑S^TM relative to our in‑house CHO‑3E7 platform—a 6‑fold increase for Depemokimab and an 11‑fold increase for Eldelumab. In contrast, Bococizumab displayed only a modest improvement, with a 1.5‑fold increase in ExpiCHO‑S^TM (Figure 4, right), reinforcing that performance gains remain target‑specific.

Building on this, we expressed Sacituzumab using the Thermo MAX protocol and purified it under low‑endotoxin conditions via Protein A affinity chromatography. The resulting high‑yield, clean preparation further strengthened our confidence in ExpiCHO‑S^TM as a robust platform for recombinant antibody production (Figure 4, left).

In Summary

Our evaluation has shown that the ExpiCHO‑S^TM cell line can have clear advantages for high yielding expression of secreted protein targets, especially antibodies, and is a valuable addition to our mammalian host cell portfolio that we offer.

At Sygnature Discovery, we work closely with clients to identify the most suitable expression strategy for each target and project, supporting efficient and scalable routes to recombinant protein production.