Cytiva ÄKTA™ pure 25M Protein Purification System

Overview

The Cytiva ÄKTA™ pure 25M is a modular, benchtop protein purification system engineered for precision, reproducibility, and method scalability in academic, biopharmaceutical development, and QC laboratories. It operates on the fundamental principles of liquid chromatography—specifically gel filtration chromatography (GFC), ion exchange (IEX), hydrophobic interaction (HIC), and affinity chromatography—enabling high-resolution separation of proteins, antibodies, viral vectors, and other biomolecules based on size, charge, hydrophobicity, or ligand-specific binding. The system integrates a high-pressure, low-pulsation dual-plunger pump, real-time multi-parameter monitoring (UV absorbance, conductivity, pH), and robust fluidic architecture capable of sustained operation at up to 20 MPa—supporting both analytical-scale characterization and preparative purification (up to 25 mL/min nominal flow, extendable to 50 mL/min during column packing). Its design conforms to Good Laboratory Practice (GLP) and Good Manufacturing Practice (GMP)-aligned workflows, with hardware and software features supporting audit trails, electronic signatures, and data integrity requirements under FDA 21 CFR Part 11.

Key Features

• Modular hardware platform allowing configuration for GFC, IEX, HIC, affinity, and reversed-phase applications—validated for use with Cytiva’s prepacked columns (e.g., HiLoad®, Tricorn™, XK™ series) and custom-packed media.
• Multi-wavelength UV detection (190–700 nm) enabling simultaneous monitoring of aromatic amino acids (280 nm), nucleic acid contaminants (260 nm), and broad-spectrum spectral scanning for peak identification and purity assessment.
• Integrated conductivity and pH sensors with calibrated, temperature-compensated output—essential for gradient elution control, buffer exchange verification, and process consistency across batches.
• High-accuracy flow delivery (±1.2% accuracy, RSD <0.5% under standard operating conditions), ensuring reproducible retention times and fraction collection across repeated runs.
• Chemical-resistant fluid path constructed from PEEK, titanium, and sapphire components—validated for daily cleaning-in-place (CIP) with 1 mol/L NaOH and compatibility with acetonitrile, isopropanol, and other organic modifiers used in reversed-phase peptide purification.
• Expandable valve manifold supporting up to 10 column positions—enabling automated multi-step purification protocols (e.g., capture → intermediate wash → polishing) without manual intervention.

Sample Compatibility & Compliance

The ÄKTA™ pure 25M accommodates sample volumes from 10 µL to >100 mL and handles viscous or particulate-containing lysates when paired with appropriate in-line filtration (e.g., 0.45 µm or 0.22 µm filters). It supports native and denaturing conditions, including low-pH elution for monoclonal antibodies and high-salt buffers for HIC. All critical operational parameters—including flow rate, pressure, UV absorbance, conductivity, and pH—are logged with timestamped metadata in UNICORN software, fulfilling traceability requirements per ISO/IEC 17025, USP , and ICH Q5A/Q5D. The system is CE-marked and compliant with IEC 61010-1 for laboratory equipment safety.

Software & Data Management

Controlled exclusively via UNICORN 7 software—a validated, Windows-based platform supporting method creation through drag-and-drop templates, real-time chromatogram overlay, and automated fraction triggering based on UV threshold, volume, or time windows. UNICORN provides full audit trail functionality (user login, parameter changes, run start/stop events), electronic signature capability, and export options compliant with CSV, PDF/A, and XML formats for regulatory submission. Raw data files (.chm, .frt) are stored in a structured, non-proprietary directory hierarchy, facilitating integration with LIMS and ELN systems. Optional UNICORN Connect enables remote monitoring and secure cloud backup (GDPR- and HIPAA-aligned configurations available).

Applications

• Purification of recombinant proteins (e.g., His-tagged, GST-fusion) using immobilized metal affinity chromatography (IMAC) or glutathione resin.
• Buffer exchange and desalting of labile enzymes prior to structural analysis (X-ray crystallography, cryo-EM).
• Polishing steps for therapeutic monoclonal antibodies—including removal of aggregates (GFC), host cell proteins (IEX), and leached Protein A (affinity).
• Characterization of viral vector purity (AAV, lentivirus) via size-exclusion chromatography coupled with multi-angle light scattering (SEC-MALS) using optional external detectors.
• Process development studies requiring method transfer between ÄKTA™ pure 25M (lab scale) and ÄKTA™ avant 150 (pilot scale), leveraging identical UNICORN methods and column geometry scaling rules.

FAQ

Can the ÄKTA™ pure 25M be upgraded to support dual-wavelength UV detection?

Yes—existing ÄKTA™ pure L systems can be retrofitted with the U9-T UV monitor to enable simultaneous 260 nm and 280 nm detection. The 25M model ships with the multi-wavelength U9-M detector as standard.
Is method validation support available for GMP environments?

Cytiva provides IQ/OQ documentation packages, UNICORN 7 validation templates, and on-site qualification services aligned with ASTM E2500 and Annex 11 requirements.
What column formats are natively supported?

Tricorn™ 5/10/20, HiTrap™, HiPrep™, and XK™ series columns are fully auto-configured in UNICORN; custom columns may be defined manually with user-specified dimensions and flow limits.
Does the system support gradient programming beyond linear profiles?

UNICORN 7 supports linear, concave, convex, step, and user-defined custom gradients—each programmable with independent dwell volume compensation.
How is system sanitization performed between runs?

Automated CIP sequences—using NaOH, ethanol, or acetonitrile—are programmable within UNICORN and monitored via conductivity/pH feedback to ensure endpoint confirmation.