Ion Exchange Chromatography

Introduction

Q, DEAE and CM Ceramic HyperD® F ion exchange chromatography  resins are designed using proprietary 'gel-in-a-shell' design that provides rapid mass transfer and effective binding. It relies on a high capacity hydrogel, polymerized within the gigapores of a rigid ceramic bead.

Features and Benefits

• High dynamic binding capacity at high flow rates
• Truly rigid, non-compressible resin
• Salt tolerant CM Ceramic HyperD® F resin reduces Ultrafiltration/Diafiltration (UF/DF) requirements

• Recombinant proteins
• Plasmid purification
• Purification of enzymes, plasma proteins
• Vaccines
• Direct capture of monoclonal antibodies (CM Ceramic HyperD® F)
• IgM

* BSA = Bovine Serum Albumin
¹ 5 mg/mL BSA in 50 mM Tris-HCl, pH 8.6
²5 mg/mL hu IgG in 50 mM sodium acetate, 100 mM NaCl, pH 4.7

Introduction

Q, DEAE and CM Ceramic HyperD® F ion exchange chromatography  resins are designed using proprietary 'gel-in-a-shell' design that provides rapid mass transfer and effective binding. It relies on a high capacity hydrogel, polymerized within the gigapores of a rigid ceramic bead.

Features and Benefits

• High dynamic binding capacity at high flow rates
• Truly rigid, non-compressible sorbent
• Salt tolerant CM Ceramic HyperD F sorbent reduces Ultrafiltration/Diafiltration (UF/DF) requirements

• Recombinant proteins
• Plasmid purification
• Purification of enzymes, plasma proteins
• Vaccines
• Direct capture of monoclonal antibodies (CM Ceramic HyperD® F)
• IgM

* BSA = Bovine Serum Albumin
¹ 5 mg/mL BSA in 50 mM Tris-HCl, pH 8.6
²5 mg/mL hu IgG in 50 mM sodium acetate, 100 mM NaCl, pH 4.7

Introduction

Q, DEAE and CM Ceramic HyperD® F ion exchange chromatography  resins are designed using proprietary 'gel-in-a-shell' design that provides rapid mass transfer and effective binding. It relies on a high capacity hydrogel, polymerized within the gigapores of a rigid ceramic bead.

Features and Benefits

• High dynamic binding capacity at high flow rates
• Truly rigid, non-compressible resin
• Salt tolerant CM Ceramic HyperD® F resin reduces Ultrafiltration/Diafiltration (UF/DF) requirements

• Recombinant proteins
• Plasmid purification
• Purification of enzymes, plasma proteins
• Vaccines
• Direct capture of monoclonal antibodies (CM Ceramic HyperD® F)
• IgM

* BSA = Bovine Serum Albumin
¹ 5 mg/mL BSA in 50 mM Tris-HCl, pH 8.6
²5 mg/mL hu IgG in 50 mM sodium acetate, 100 mM NaCl, pH 4.7

Salt Tolerant Advanced Recovery Anion Exchange Chromatography Resin

HyperCel STAR AX resin is an industry-scalable anion exchange chromatography resin designed for high productivity protein capture at moderate or higher salt conductivity (2 to 15 mS/cm), typical of undiluted biological feedstocks (i.e., mammalian cell culture supernatants, E. coli feedstock, plasma, others…).

Applications include direct capture of recombinant proteins, monoclonal and polyclonal antibodies, plasma derivatives or other biopharmaceuticals.

Due to its ability to capture directly proteins from undiluted feedstreams, HyperCel STAR AX resin can also be used for early contaminant removal (i.e., CHO Host Cell Proteins), before target purification, i.e., before MAb capture by a Protein A affinity step.

HyperCel STAR AX resin is manufactured at large scale and meets the needs of industrial users and regulatory authorities. This resin offers:

• High dynamic binding capacity (DBC) at short residence time (2 minutes or lower)
• Direct capture of protein from undiluted feedstock at moderate or high conductivity
• Excellent flow rate properties for fast feedstock processing
• Distinctive selectivity, consistent in a broad conductivity range (2 – 15 mS/cm)
• Enhanced process economics

Application 1. Direct Capture of Albumin from Undiluted Plasma

The objective is to evaluate HyperCel STAR AX resin as the first step in a two-step purification sequence to capture human serum albumin (HSA) from undiluted plasma (conductivity 11 mS/cm). HyperCel STAR AX resin was used as a first capture step, followed by an orthogonal cation exchange step, without pH or conductivity adjustment of the plasma. Neat undiluted plasma (pH 7.6, 11mS/cm), was loaded with DBC of 30 mg/mL.

Design of Experiments (DoE) in 96-well filter plates was performed to determine optimal conditions to achieve the best yield/purity ratio.

These conditions were then transferred to column chromatography on PRC prepacked columns.

Conditions determined in 96-well plates were applied to chromatography of undiluted plasma on HyperCel STAR AX resin in a 1 mL PRC prepacked column. Chromatogram and SDS-PAGE analysis of fractions confirmed that most contaminants were eliminated by high conductivity wash, leading to a 99% pure HSA fraction in a single step, with 90% yield.

In addition, HyperCel STAR AX resin used at capture step with undiluted plasma had a DBC >30 mg/mL, more than 2-fold higher than that of a conventional DEAE agarose resin tested in these conditions.

HSA was eluted by a simple decrease of pH (4.0), without addition of salt, allowing a direct load on an orthogonal cation exchange column.

Application 2. Early Removal of CHOPs before Protein A Capture of a Monoclonal Antibody (MAb)

The objective of this study was to evaluate the impact of a pre-purification step using HyperCel STAR AX resin before conventional Protein A resin capture of a monoclonal antibody (MAb) from mammalian cell culture feedstock.

The content of contaminating CHOPs (Chinese Hamster Ovary Proteins) was compared using a commercial ELISA assay for the two purification schemes.

Data shows that using HyperCel STAR AX resin prior to Protein A results in a better CHOP reduction (>8-fold). Due to this synergistic effect, starting from an initial Host Cell Protein (HCP) content of 413,000 ppm, the final CHOP level is reduced to ∼400 ppm (3-Log reduction), with a MAb recovery of 90%.

A pre-purification step can impact positively process economics by extending the lifetime of an expensive Protein A column used as standard step for MAb capture.

HyperCel STAR AX resin is composed of a rigid cellulose matrix that has excellent flow properties and generates low backpressure, compatible with the needs of manufacturing-scale protein production. The resin is available in a variety of packaging configurations as well as convenient 1 mL and 5 mL PRC prepacked columns designed for fast method optimization, selectivity screening or small preparative work. HyperCel STAR AX resin is supplied as a slurry/suspension in 1 M NaCl containing 20% (v/v) ethanol or as a moist cake for process-scale applications (the moist cake resin facilitates the resin transfer, avoiding the agitation and suspension of large material volumes).

HyperCel STAR AX resin has a chemical stability that ensures simple clean-in-place (CIP) and storage. For standard CIP, 0.5 to 1 M NaOH treatment is recommended, while long-term storage in 10 to 100 mM NaOH is possible.

Main Properties

¹ Determined using a 5 mg/mL BSA in 25 mM Tris-HCl , 0.14 M NaCl at 2 minute residence time.

² Injection of 5 column volumes (CV) of 0.5 – 1 M NaOH, 1 hour contact time.

HyperCel STAR AX Resin

HyperCel STAR AX PRC Prepacked Columns