Cytokines and growth factors: what are international units?

In cell biology, immunology, and drug development, cytokines and growth factors are rarely discussed purely in terms of mass. Instead, their potency is often expressed in International Units (IU) a convention that can confuse even experienced researchers. Why are some cytokines defined in IU, others sold only by weight, and why do IU values seem to vary between suppliers?

Understanding the rationale behind IU, their inherent variability, and their limited applicability is essential for reproducible science and informed experimental design.

What Is an International Unit?

An International Unit (IU) is a measure of biological activity, not physical quantity. Unlike milligrams or micrograms, which measure how much protein is present, IU measure what the protein does in a defined biological system.

Formally, an IU is defined by comparison to a reference standard, typically established by the World Health Organization (WHO). For a given cytokine or growth factor, a WHO International Standard is assigned an arbitrary but fixed number of IU. All other preparations are then calibrated against this standard using a defined biological assay.

In simple terms:

1 IU equals the biological activity of 1 IU of the reference standard under defined assay conditions.

There is no universal mass equivalent for an IU.

Why IU Are Used for Cytokines and Growth Factors

Cytokines and growth factors are biological signals, not just chemicals. Their functional impact depends on more than how much protein is present. Factors such as protein folding, post-translational modifications, aggregation, oxidation, and degradation can dramatically affect activity (up to several fold) without changing mass. In other words, two cytokine samples with identical weights can differ several-fold in biological potency.

Using IU allows researchers to:

• Compare activity across batches and manufacturers
• Normalize dosing to biological effect rather than protein quantity
• Account for structural or functional differences between preparations

This is particularly important for cytokines such as IL-2, interferons, GM-CSF, and G-CSF, where downstream cellular responses are highly sensitive to potency.

How IU Are Determined

IU assignment is based on bioassays, not chemical analysis. Each cytokine or growth factor has one or more historically accepted assays that reflect its primary biological function.

Examples include:

• IL-2: T-cell proliferation (typically assayed using CTLL-2 cells)
• Interferons: Antiviral protection assays
• Colony-stimulating factors: Hematopoietic cell proliferation or differentiation assays
• EGF or FGF: Mitogenic assays in responsive cell lines

A test sample and the reference standard are run side by side, dose–response curves are generated, and activity is assigned by relative potency. This means IU are comparative by definition, not absolute.

Why IU Values Vary

Even when traceable to a WHO standard, IU values generated by the same cytokine sample assayed in different labs are inherently variable. This variability arises from several sources:

1. Biological Assay Variability

Bioassays involve living cells, which are influenced by:

• Cell passage number
• Culture conditions
• Serum components
• Readout method (radioactive, colorimetric, luminescent)
• Curve-fitting models

Small differences can lead to measurable shifts in calculated activity.

2. Protein Heterogeneity

Recombinant cytokines may differ in:

• Glycosylation (especially mammalian vs bacterial expression), although this is not crtitcal for many cytokines and applications
• N-terminal processing
• Oxidation state
• Aggregation level

These differences can affect receptor binding and signaling efficiency.

3. Stability Over Time

Bioactivity can decrease during storage or handling while mass remains unchanged. IU therefore represent activity at the time of testing, not an immutable property.

Because of these factors, IU values are best understood as approximations with traceability, not exact constants.

Why Not All Growth Factors Have IU Standards

Despite their usefulness, IU are not universally applicable, and many growth factors do not have WHO-assigned IU standards. There are several reasons for this.

1. Lack of a Clear, Consensus Bioassay

Some growth factors act on multiple cell types or pathways, making it difficult to define a single, representative biological assay. Without a widely accepted assay, an IU standard cannot be meaningfully established.

2. Limited Clinical or Regulatory Need

IU standards are most often developed for proteins with:

• Clinical use
• Regulatory relevance
• Broad international distribution

Many growth factors are primarily research tools, and the cost and effort of establishing a formal international standard may not be justified.

3. Context-Dependent Biology

Some growth factors exhibit context-specific effects that vary dramatically with cell type, receptor expression, or co-factors. In such cases, a single IU definition may be misleading rather than helpful.

4. Historical and Practical Considerations

IU standards have accumulated over decades based on scientific priority and clinical demand. Their absence does not imply lesser importance—only that standardization has not occurred.

Why Companies Sell Growth Factors by Weight, Not IU

Even when IU exist, most suppliers sell cytokines and growth factors by mass, reporting IU only as a characterization metric (e.g., IU/mg).

This reflects a fundamental distinction:

• Mass is absolute, traceable, and verifiable
• IU are assay-dependent and biological

Selling by weight minimizes regulatory complexity and liability while allowing users to calculate approximate activity using the reported specific activity.

Practical Takeaways for Researchers

• IU measure biological potency, not amount
• IU are meaningful only in the context of a defined assay and reference standard
• Variability in IU is inherent and unavoidable
• Not all growth factors have IU because not all can be standardized biologically
• Always consult specific activity (IU/mg) and assay descriptions when comparing products

Conclusion

International Units are a powerful tool for expressing the biological activity of cytokines and growth factors, but they are not universal, absolute, or immutable. They reflect a compromise between biological relevance and experimental variability. Understanding their limitations—and why they exist at all—allows researchers to design better experiments, interpret data more accurately, and avoid false assumptions about equivalence between reagents.

In the end, growth factors are not just molecules; they are signals. IU exist to remind us of that distinction.

IMAGE Understanding IUs CREDIT CellGS

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