Clearing the Confusion: GPC, SEC, GFC –What , When, Why, and How?

Importance of the topic

Size‐based separation techniques such as gel permeation chromatography (GPC), size exclusion chromatography (SEC) and gel filtration chromatography (GFC) are essential tools in polymer science and biopharmaceutical analysis. They provide molecular size distribution profiles, enabling critical insights into polymer polydispersity, biopolymer aggregation and purity. Accurate size separation underpins quality control, research and development, and regulatory compliance in both industrial and academic laboratories.

Objectives and overview of the presentation

The source document aims to:

• Clarify terminology around GPC, SEC and GFC.
• Describe when and why to choose size‐exclusion techniques for polymers versus biomolecules.
• Outline practical considerations for column selection, eluents, calibrants and method development.

Methodology and used instrumentation

GPC/SEC separations employ columns packed with porous beads. Molecules in dilute solution elute based on hydrodynamic volume: larger species elute first, smaller species interact more deeply with pores and elute later. Key method steps include sample dissolution, injection, separation and detection. Instruments and consumables discussed include:

• SEC columns with various chemistries (polymethacrylate, divinylbenzene, silica‐diol).
• Column dimensions and pore sizes tailored to molecular weight ranges (e.g. 300 Å for mAb analysis).
• Detectors such as differential refractive index (DRI), UV and light scattering for molecular weight measurement.
• Eluent delivery systems with temperature control and filtering to maintain separation integrity.

Main results and discussion

Key points from the review:

• Nomenclature: GPC, SEC and GFC are synonymous modes distinguished by application context (polymers vs biomolecules).
• Hydrodynamic volume governs separation, not absolute molecular weight—different polymers of equal mass may elute differently.
• Pore size selection is critical: too small leads to exclusion; too large reduces resolution. Combining columns in series extends molecular weight range and improves separation.
• Mobile phase choice must suppress non‐specific interactions: organic solvents with salts (e.g. THF + salt) for polymers; buffered aqueous phases (e.g. 150 mM phosphate, pH 7) for proteins.
• Calibration standards need to match sample chemistry and solvent to yield reliable molecular weight distribution data.

Benefits and practical applications

Size‐exclusion chromatography provides:

• Detailed distribution profiles, not just averages, enabling prediction of polymer processing behavior and material properties.
• Quantitative analysis of protein monomer content and aggregate levels critical to biopharmaceutical safety and efficacy.
• High reproducibility for routine quality control in polymer manufacturing and therapeutic protein production.

Future trends and possibilities

Emerging directions include integrating multi‐detector systems (e.g. light scattering, viscometry), advanced column materials with narrower pore distributions, and automated data analysis workflows. Coupling SEC with mass spectrometry for deeper structural characterization and applying microfluidic SEC formats for high‐throughput screening are promising avenues.

Conclusion

GPC/SEC remains the only common technique to resolve molecular size distributions, offering indispensable insights in polymer science and biopharma. Proper column selection, mobile phase optimization and calibration are keys to robust, reproducible results. Continued innovation in column chemistries, detection methods and automation will expand the technique’s capabilities and applications.