Ultrasensitive method for profiling host cell proteins by coupling limited digestion to ProteoMiner technology

Area of Science:

• Biopharmaceutical analysis
• Analytical chemistry
• Protein biochemistry

Background:

• Host cell proteins (HCPs) are critical process-related impurities in therapeutic proteins.
• Monitoring HCPs is essential for ensuring drug product quality and patient safety.
• Low abundance HCPs pose a significant challenge for detection using liquid chromatography-mass spectrometry (LC-MS).

Purpose of the Study:

• To improve the enrichment of low-abundance host cell proteins (HCPs) in therapeutic protein products.
• To enhance the sensitivity of HCP detection using mass spectrometry.
• To establish a more comprehensive HCP profiling method for biopharmaceutical quality control.

Main Methods:

• Coupling limited digestion with ProteoMiner technology (PMLD) for HCP enrichment.
• Utilizing liquid chromatography coupled to mass spectrometry (LC-MS) for high-sensitivity detection.
• Analyzing a NIST monoclonal antibody preparation to validate the method.

Main Results:

• Achieved a 7694-fold enrichment of low-abundance HCPs.
• Enabled detection of HCPs at concentrations as low as 0.002 ppm.
• Identified 850 HCPs with high confidence, a 40% increase over previous methods.

Conclusions:

• The PMLD technique significantly enhances HCP enrichment and detection sensitivity.
• This improved method allows for more comprehensive HCP profiling in biotherapeutics.
• The findings contribute to more robust quality control strategies for therapeutic protein products.