Pharma Biologics

Protein-based biologics

Biopharmaceutical proteins, such as monoclonal antibodies (mAbs), fusion proteins, and other protein-based drugs, are highly sensitive molecules.

During manufacturing and handling, they face multiple stressors that can induce aggregation, impacting yield, efficacy, and patient safety.

Detecting and controlling aggregates – from early-stage development to fill-finish – remains a persistent challenge in pharma biologics.

Limitations in Protein Aggregate Monitoring

• Unexpected aggregate formation during pumping, mixing, or filtration?
• Difficulty tracking both nanoscale and subvisible particles in one run?
• Incomplete process insight due to reliance on separate, offline instruments?
• Aggregates forming too late in DSP to intervene?
• Protein destabilization without clear root causes?

These issues are well-recognized across the industry – and solvable with the right tools.

The NanoFlowSizer offers a novel dual-mode solution for real-time monitoring of protein aggregation: inline, at-line, or offline.

• LPD (Large Particle Detection): provides imaging-based detection of rare, subvisible aggregates >1 µm at concentrations down to 10² particles/mL.
• PhaSR-DLS (Phase-sensitive Dynamic Light Scattering): detects monomers and submicron aggregates with high sensitivity and noise suppression — even at low concentrations.

Together, they deliver spatially resolved, real-time data on aggregation state across a wide range of concentrations and size distributions — using non-invasive optics and without sampling or dilution.

In a representative downstream model, BSA protein solutions were recirculated through a peristaltic pump to simulate shear stress commonly encountered in biopharmaceutical processing. The NanoFlowSizer, equipped with both PhaSR-DLS and LPD, continuously monitored aggregation during this process.
Results showed a near-linear increase in both submicron (150–400 nm) and subvisible (>1 µm) aggregates over time. These populations, traditionally requiring separate instruments to detect, were measured simultaneously using a single system.

“The unique combination of PhaSR-DLS and LPD in the NanoFlowSizer effectively tracks both submicron and rare subvisible protein aggregates… The method can advance protein DSP by reducing complexity, costs, and boosting efficiency, while safeguarding product quality.”

This real-time, non-invasive insight into mechanical stress-induced aggregation offers biologics manufacturers a practical solution for better risk mitigation, quality assurance, and process control.

Using Bovine Serum Albumin (BSA) as a model protein, the graph shows the real-time change in the hydrodynamic diameter (Zaverage) of the BSA/PEI complex as varying amounts of BSA are added. Different ratios of BSA-to-PEI ratios were mixed with 250 kDa PEI in dilute PBS (pH 7.4, isotonic ionic strength, total sample volume of 2 mL).

Large Particle Detection (LPD)

• Detects rare particles >1 µm using imaging with lateral scanning
• Capable of measuring particle concentrations <10³ particles/mL
• Provides real-time visualization and size statistics on large aggregates

PhaSR-DLS (Phase-sensitive Spatially Resolved DLS)

• Measures down to <10 nm, with enhanced sensitivity for dilute samples
• Eliminates baseline distortion from number fluctuations
• Suitable for proteins, mAbs, LNPs, and nanocarriers in complex matrices

Measurement Benefits

• Non-invasive, no sampling or dilution
• Inline or at-line, real-time measurements
• Broad size range: <10 nm to >10 µm
• Compatible with a wide range of container types (syringes, IV bags, process vessels)