This document describes the separation of recombinant human erythropoietin (rEPO) using Agilent Bio SEC-3. The method utilizes Agilent Bio SEC columns, which are designed to improve separation efficiency and stability by reducing secondary interactions. The columns are optimized for use with organic modifier denaturants and low salt concentrations, making them ideal for quality control in protein primary structure analysis.
The document details a method for monitoring the purity and stability of recombinant human Erythropoietin (rEPO). It utilizes size exclusion chromatography with Agilent Bio SEC-3 columns to separate intact rEPO from aggregates, dimers, and monomers. The study also evaluates the separation performance under stress conditions, specifically heat-stressed samples at low pH and neutral pH.
| Column Part Number | 5190-250 |
|---|---|
| Column Dimensions | 4.6 x 300 mm |
| Particle Size | 3 um |
| Flow Rate | 0.35 mL/min |
| Injection Volume | 5 ul |
| Mobile Phase | 150 mM sodium phosphate buffer, pH 7.0 |
| Detector Wavelength | 225 nm |
The Agilent 1260 Infinity Bio-inert Quaternary LC system.
The optimized flow rate was 0.35 mL/min.