In the fast-paced world of biologics development, analytical assays play a pivotal role in ensuring product safety, efficacy, and regulatory compliance. Yet, despite technological advances, some assays continue to pose significant challenges across the product lifecycle. In a recent conversation with Dr Nadine Ritter, a leading expert in analytical sciences for biological products, two particular assays stood out as the most analytically and strategically complex: host cell protein (HCP) assays and potency assays.

The Hidden Complexity of Host Cell Protein Assays

Every biologic, by definition, is derived from a living system—whether it be CHO cells, E. coli, yeast, or other expression systems. These cells don’t just produce the target therapeutic protein; they also generate a vast array of other proteins necessary for their survival and function. The downstream purification process aims to remove these unwanted proteins, yet complete removal is virtually impossible. Residual host cell proteins remain, and therein lies the analytical challenge.

Dr Ritter explains that unlike DNA, which can be addressed using relatively standardised platform methods due to the consistency of the genome, host cell proteins represent the cell’s proteome—a far more variable and complex landscape. Factors such as cell line, age, culture conditions, and even the phase of cell death can affect the protein expression profile, creating a potential pool of thousands of unique contaminants.

These residual proteins are not only analytically elusive—often present below the detection limits of standard assays—but also potentially problematic. Some may elicit immune responses in patients due to the body’s innate sensitivity to foreign proteins. Others may silently catalyse degradation pathways in the final product, compromising stability and quality over time.

To meet this challenge, the industry has embraced tools like 2D-DIGE (two-dimensional differential gel electrophoresis) and mass spectrometry, which allow for deeper characterisation of HCP profiles. Meanwhile, the advent of commercially available HCP ELISAs has filled a critical gap, enabling earlier-stage monitoring before process-specific assays can be developed.

However, Dr Ritter cautions against complacency. “There’s so much misinformation about quantitative techniques and what you see or don’t see,” she warns. Without a robust HCP characterisation and control strategy, developers risk making decisions based on incomplete data—decisions that can reverberate throughout the development process.

The Potency Puzzle: Building Reliable Functional Assays

If HCP assays represent a challenge in detection and characterisation, potency assays raise the stakes even further by requiring a consistent and legally mandated measure of biological activity. Potency is not just a desirable attribute—it’s a statutory requirement. It differentiates a biologically active molecule from one that may appear identical on paper but lacks therapeutic effect.

Many potency assays rely on relative activity measurements, meaning they compare a test sample to a well-characterised reference standard. And herein lies the conundrum: when developing a new biological product, that reference doesn’t exist yet. The very first batch chosen to serve as the potency reference becomes the foundational standard for all future assays—potentially all the way from early clinical trials through to commercialisation.

“It’s literally a chicken-and-egg scenario,” says Dr Ritter. Early choices around the first reference standard—its stability, characterisation, and how well it reflects the product’s intended function—can lock developers into a trajectory that’s difficult and costly to adjust later. Transitioning from one reference standard to another is particularly difficult in potency assays, far more so than in physical assays.

Despite these challenges, the field has made significant progress. Critical reagents are now better characterised, and ready-to-use cell lines have become more common, reducing variability in assay performance. But according to Dr Ritter, the biggest challenge isn’t the assay itself—it’s the control and continuity of the reference standard strategy that underpins it.

A Regulatory Grey Area

One of the most striking observations Dr Ritter shared is the lack of detailed regulatory guidance on these topics. While regulatory bodies mandate the use of HCP and potency assays, their guidance often lacks the granularity practitioners need. This absence of detailed instruction contributes to confusion, inconsistent practices, and, in some cases, strategic missteps that are difficult to recover from.

“There are bits and pieces of best practices scattered across industry forums,” Ritter notes, referring to initiatives like CMC strategy meetings and conferences hosted by organisations such as CASSS. These events provide valuable case studies and community-driven insights, but they are no substitute for a clear, centralised regulatory framework.

As a result, navigating these complex assays requires more than technical skill—it demands strategic foresight and a willingness to learn from the broader scientific community. “Come to a class,” Ritter encourages, emphasising the importance of education and ongoing professional development in mastering these nuanced areas.

Final Thoughts

Analytical assays are often viewed through a technical lens, but as Dr Nadine Ritter makes clear, their implications stretch far beyond the lab bench. Host cell protein assays and potency assays each come with their own unique challenges—scientific, strategic, and regulatory—that can have long-term impacts on product development and success.

For professionals in biologics development, these insights are a reminder that early decisions matter. Investing in the right tools, understanding the limitations of your assays, and planning for long-term assay control are not just best practices—they’re essential safeguards for ensuring product quality, regulatory compliance, and, ultimately, patient safety.

Watch the full interview below:

Written by Educo Life Sciences Expert, Nadine Ritter

Dr Nadine M. Ritter is a renowned expert in analytical sciences for biologics, with a PhD in cell and molecular biology from Rice University. Her early academic research focused on bone mineralisation and extracellular matrix proteins. She began her industry career at Abbott Laboratories, where she specialised in protein chemistry, analytical method development, and regulatory compliance. As Director of Analytical Services at BioReliance, she led CMC teams supporting biologics characterisation, QC testing, and regulatory submissions. In 1999, Dr Ritter pioneered public training on biologic stability programmes, evolving into comprehensive CMC analytics courses. Since 2002, she has worked as a global consultant and educator, contributing to over 120 international IND/IMPD and BLA/MAA filings. Through her company, Global Biotech Experts, LLC, she collaborates with elite consultants to support GMP, PAI, and analytical challenges. Dr Ritter remains a sought-after speaker, trainer, and CMC advisor to leading biopharma companies worldwide.

This article was written from an interview (which you can watch above)