Comparison of purification performance of Amsphere A3 to other commercially available Protein A resins
Posted on August 22,2017
Protein A affinity chromatography represents the key initial capture method in monoclonal antibody (mAb) purification. There are three major types of protein A “resin” based on a different matrix chemistries – glass, agarose and synthetic polymer. State of the art resins must offer good specificity, high mass transfer and binding capacity, low non-specific adsorption, low ligand leakage, suitable back pressure under high flow operation, and good chemical stability – particularly during alkaline sanitization.
Until now, selecting a protein A resin for bioprocessing applications involved balancing among high specificity, high mass transfer and binding capacity, low non-specific adsorption and ligand leakage, incompressibility, resistance to alkaline condition for sanitization, chemical stability and cost effectiveness. Recent results using new Amsphere A3 versus Polymer H and Agarose show that the Amsphere A3 design minimizes compromise in performance criteria.
TABLE 1: HOST CELL PROTEIN LEVEL OF LOADING SAMPLES
Amsphere A3 is a new protein A resin designed with a surface modified base bead and alkali-resistant optimized ligand.
Protein A ligand
• High DBC via controlled conformation and orientation
• High alkaline stability from protein engineering Surface modification
• Low HCP levels by surface hydrophilization Base bead formulation
• High DBC at high flow rate
• Excellent pressure and flow properties via rigid crosslinking
FIGURE 1: TEST CONDITIONS OF CHROMATOGRAPHY PROCESS
Study Methods and Materials
Polymer (H) protein A affinity chromatography resins were tested in comparison with Amsphere A3. Five mAb samples (as in Table 1) expressed in CHO cells and clarified by centrifugation and 0.22 mm filtration were tested.. Purified polyclonal human IgG was also used. Dynamic binding capacity (DBC) at 10% breakthrough versus residence time, alkali stability (0.5 M NaOH), host cell protein (HCP) clearance, pressure-flow properties, and protein A ligand leaching were evaluated according to common literature protocols.
The details of the protein A column chromatography processes are given in Figure 1. Other experimental details are given in association with the results below.
1. DYNAMIC BINDING CAPACITY
2. mAbs DYNAMIC BINDING CAPACITIES AT 5 MINUTES RESIDENCE TIME
3. HCP CLEARANCE – TARGET POOL HCP LEVELS
4. LEACHED PROTEIN A INTO TARGET POOL
5. ALKALINE STABILITY (% RETENTION OF DBC) TO 0.5M NaOH
6. PRESSURE-FLOW PROPERTIES
Amsphere A3 has incorporated many resin design attributes that minimize compromise in key selection criteria such as DBC, HCP reduction, and caustic stability. The selection of ligand design, bead design and surface modification result in a superior product, fit for a wide range of operating conditions and scale.
AmsphereTM is a global trademark of JSR Corporation
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