High-Performance Liquid Chromatography (HPLC)

Chromotography Services for Biologics:Gene Therapy, mRNA, LNPs & ADCs

GMP-Compliant HPLC Testing for IND Support, Product Characterization, and Release

BioAnalysis provides a complete suite of GMP-compliant HPLC services for biologics developers working on gene therapies, cell therapies, mRNA vaccines, lipid nanoparticles (LNPs), monoclonal antibodies (mAbs), and antibody-drug conjugates (ADCs). Our HPLC testing approach supports method development, product characterization, stability testing, comparability studies, and GMP lot release—with capabilities aligned to regulatory expectations.

High-Performance Liquid Chromatography (HPLC) is critical for characterization, identification, quantification of drug product, and excipients, as well as purity/impurity limit testing.

Our expert staff have developed sensitive, specific, and robust bespoke assays for residuals, stability testing and excipients. We thrive on the most challenging product identification and characterization.

We take advantage of a broad range of detectors including charged aerosol detection (CAD), ultraviolet/visible (UV/VIS), fluorescence, and size exclusion chromatography-multi-angle light scattering (SEC-MALS).

We specialize in biologics with expertise in Size Exclusion Chromatography (SEC), Reverse Phase (RP), Ion Exchange (IEX), and Hydrophilic Interaction (HILIC).

HPLC separates molecules based on hydrophobicity—ideal for peptides, small molecules, and impurities.  For complex biologicals, RP is critical to verify product identity, purity, and stability. RP-HPLC is the cornerstone of qualification and identity testing methods for Chemistry, Manufacturing, and Controls (CMC) packages. 

  • Applications:
    • Residual solvent and impurity analysis and limit testing
      • Transfection reagents: FectoVIR®, Polyethylenimine (PEI), PEIpro®, TransIT-VirusGEN®, RevIT™,
      • Detergents/Surfactants: Polysorbates (Tween) P80,  P20, Poloxamer P188 (Pluronic F-68), Triton X-100
      • Purification reagents: CsCl, iodixanol, affinity ligands, polyethylene glycol (PEG)
    • Excipients content and quantification
      • Sorbitol, Polysorbates (Tween) P80 or P20, Poloxamer P188
    • mRNA concentration in LNPs
    • Lipid content
    • Verification of peptide mapping and sequence
    • Detection of degradation products in mRNA, AAVs, or ADCs
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Size Exclusion Chromatography (SEC)

SEC analysis separates biomolecules based on size to detect aggregates, fragments, and oligomers.

Aggregation is a critical impurity that can affect efficacy, safety, and immunogenicity. Regulatory agencies require evidence that aggregates are well controlled and SEC helps de-risk therapeutics in development by detecting unwanted size variants early.

  • Applications:
    • Release and stability testing for all biologics including mAb/ADC/AAV vectors
    • LNP particle uniformity
    • Ribonucleoprotein (RNP) complex formation

SEC-MALS (Multi-Angle Light Scattering Coupled to SEC)

SEC-MALS measures the absolute molecular weight and oligomeric state of analytes without relying on standards. 

This approach confirms that a biologic drug has the correct size and structure, especially critical for viral vectors and nanoparticle-based drugs. 

BioAnalysis SEC-MALS offers regulatory-grade data that supports product consistency testing across batches. This methodology is commonly used as a quick in-process sample testing method (non-GMP) for an estimation of empty/full capsids, capsid titer, and vector titer in one run. 
For measurement of empty/partially full/full capsids, including GMP measurement, BioAnalysis recommends our unique Analytical Ultracentrifugation testing.

  • Applications:
    • Determining molar mass of AAV capsids
    • Capsid titer and vector titer of AAVs
    • Assessing payload loading in LNPs
    • Confirming oligomeric state of protein therapeutics

Ion Exchange Chromatography (IEX – AEX + CEX)

IEX separates biomolecules by charge, AEX by negative surface charges and CEX by positive surface charges.  While AEX is ideal for detecting charge variants and nucleic acid impurities, CEX resolves basic charge variants that arise from modifications such as deamidation or lysine clipping.  Charge heterogeneity can impact potency, pharmacokinetics, and safety.  Characterizing charge also helps establish structure function relationships.

IEX methodology can detect and quantify product- and process-related impurities and ensure drug substance consistency across development and scale-up.

  • Applications:
    • Empty vs. full AAV capsid separation mainly for process support
    • mRNA or plasmid purity profiling
    • Charge isoform characterization in antibodies
    • Characterization of post-translational modifications (PTMs)

Hydrophilic Interaction Chromatography (HILIC)

HILIC separates highly polar molecules and is used primarily for glycan and polar excipient analysis.  Glycosylation is essential for monoclonal antibodies and impacts function, as well as immunogenicity.