SEC was carried out to detect monomer loss and aggregate formation (dimer, trimer and multimer) using an AKTA Pure Fast Performance Liquid Chromatography System (Cytiva). Prepacked SEC columns (Superdex 200 10/300 GL 1×30 cm, particle diameter 13 μm) with TSKgel guard column SwXL (6 mm× 40 mm) were used. MS-Hu6 samples were diluted to 1 mg/mL using the formulation buffer, loaded onto the SEC column (500 μL), and eluted isocratically at a flow rate of 0.4 mL/min. The mobile phase is composed of sodium phosphate buffer at pH 6.2, 260 mM sucrose, 0.001% w/v Tween 20, and 1 mM NaCl at 25°C. Protein concentration was measured at 280 nm. The area under absorption curve of the chromatogram was used to determine monomeric loss. The experiment was repeated twice, and representative chromatographs are reported.
Akta pure fast performance liquid chromatography system
Manufactured by Cytiva
Sourced in United States
The AKTA Pure Fast Performance Liquid Chromatography System is a laboratory instrument designed for fast and efficient separation and purification of biomolecules, such as proteins, peptides, and nucleic acids. It utilizes high-performance liquid chromatography (HPLC) technology to achieve precise and reproducible results.
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3 protocols using akta pure fast performance liquid chromatography system
1
SEC Analysis of MS-Hu6 Monomer Stability
2
Monomer Loss and Aggregation Analysis of MS-Hu6
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SEC was carried out to detect monomer loss and aggregate formation (dimer, trimer, and multimer) using an AKTA Pure Fast Performance Liquid Chromatography System (Cytiva). Prepacked SEC columns (Superdex 200 10/300 GL 1×30 cm, particle diameter 13 µm) with TSKgel guard column SwXL (6 mm× 40 mm) were used. MS-Hu6 samples were diluted to 1 mg/mL using the formulation buffer, loaded onto the SEC column (500 μL), and eluted isocratically at a flow rate of 0.4 mL/min. The mobile phase is composed of sodium phosphate buffer at pH 6.2, 260 mM sucrose, 0.001% w/v Tween 20, and 1 mM NaCl at 25 °C. Protein concentration was measured at 280 nm. The area under absorption curve of the chromatogram was used to determine monomeric loss. The experiment was repeated twice, and representative chromatographs are reported.
3
Size Exclusion Chromatography for Antibody Aggregation
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Size exclusion chromatography (SEC) was performed to detect monomer loss using an AKTA Pure Fast Performance Liquid Chromatography system (Cytiva, Marlborough, MA, USA).
Prepacked SEC columns (Superdex 200 10/300 GL 1×30 cm, particle diameter 13 µm) with TSKgel guard column SwXL (6 mm× 40 mm) were used for the analysis. SEC was used to separate the native monomeric proteins from any soluble aggregates in the antibody formulations.
Immediately before the analysis, all samples were diluted to 1 mg/ml using the formulation buffer, loaded onto the SEC column (500 µl), and eluted isocratically at a flow rate of 0.4 ml/min. The mobile phase consists of 20 mM phosphate buffer at pH 6.2, 260 mM sucrose, 0.001% w/v Tween 20, and 1 mM NaCl at 25°C. The protein concentration was measured by absorbance at 280 nm.
The area under curve of absorption peaks in the chromatogram was used to determine the % monomer loss in the antibody formulations. Representative chromatographs were reported.
Prepacked SEC columns (Superdex 200 10/300 GL 1×30 cm, particle diameter 13 µm) with TSKgel guard column SwXL (6 mm× 40 mm) were used for the analysis. SEC was used to separate the native monomeric proteins from any soluble aggregates in the antibody formulations.
Immediately before the analysis, all samples were diluted to 1 mg/ml using the formulation buffer, loaded onto the SEC column (500 µl), and eluted isocratically at a flow rate of 0.4 ml/min. The mobile phase consists of 20 mM phosphate buffer at pH 6.2, 260 mM sucrose, 0.001% w/v Tween 20, and 1 mM NaCl at 25°C. The protein concentration was measured by absorbance at 280 nm.
The area under curve of absorption peaks in the chromatogram was used to determine the % monomer loss in the antibody formulations. Representative chromatographs were reported.
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