CHT Ceramic Hydroxyapatite and Bio-Gel® Crystalline Hydroxyapatite Resins offer unique selectivities, enabling the resolution of mixtures that appear homogenous with other media. These mixed-mode resins can be used at any stage in a chromatographic workflow from initial capture to final polishing and are ideal for the purification of monoclonal antibody (mAb) products.
This whiteboard animation video provides a brief introduction to Bio-Rad’s innovative mixed-mode media, Ceramic Hydroxyapatite Resin (CHT), and walks through its unique separation properties, selectivity, and how these properties benefit downstream processing in drug development of biotherapeutics.
Advances in Separation & Purification: Purifying Monoclonal Antibodies
The affinity capture paradigm that dominates industrial IgG purification has proven unsuitable for IgMs because, in most cases, they are affected adversely by harsh elution conditions. The large size of IgMs is also a challenge because it limits the operating conditions and performance of traditional porous-particle-based chromatography media. This article describes how these challenges can be overcome with available technology to develop effective manufacturing procedures for IgM monoclonal antibodies.
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Ceramic hydroxyapatite is one of the leading media for biomolecule purification. It's a spherical ceramic form of crystalline hydroxyapatite and this form allows it to be used in production-scale columns at high flow rates while maintaining its unique separation properties.
Bio-Rad InPlace process-scale chromatography columns are designed for industrial applications with contained filling, packing, unpacking, and cleaning for safe and hygienic column operation. To learn more about Bio-Rad InPlace columns, click on the video.
CHT Packing Guidelines Video
Workflow for mAb purification with UNOsphere SUPrA affinity, UNOsphere Q, and CHT Type 1 media.
Purification of IgG using CHT Ceramic Hydroxyapatite. Read more.
A Matrix With Unique Separation Properties and Unparalleled Selectivity and Resolution CHT ceramic hydroxyapatite is a spherical, macroporous form of hydroxyapatite. The ceramic material overcomes many of the limitations of traditional crystalline hydroxyapatite and provides the throughput, stability, and reproducibility required for industrial biopharmaceutical manufacturing. It has unique separation properties and unparalleled selectivity and resolution.
Chromatogram of murine IgG1 purification on the UNOsphere S column during small-scale process development. Diluted cell culture (20 ml) was loaded onto an UNOsphere S 0.7 x 5 cm column at a flow rate of 600 cm/hr in 20 mM phosphate-citrate buffer, pH 4.0. The sample was eluted in 10 column volumes (CV) of a 0–0.5 M NaCl gradient, followed by 5 CV of 1 M NaCl in the same buffer. The column was then cleaned in 1 M NaOH. The double-headed arrow indicates the fractions (1 ml each) containing IgG1. Blue trace, A280; red trace, conductivity profile.
Plasmid Purification Using CHT Ceramic Hydroxyapatite Support
Purification of plasmid DNA on CHT II support. Buffer A: 10 mM sodium phosphate + 1 mM EDTA, pH 7.0 Buffer B: 0.4 M sodium phosphate +1 mM EDTA, pH 7.0 Flow rate: 1.5 ml/min Gradient: 0–100% buffer B for 10 column volumes Fraction size: 2.0 ml
Plamid DNA is being used successfully as a gene delivery vector in a variety of clinical application (Smith et al. 1999). Plasmids for gene therapy are usually produced in an E.coli host. One of the technical challenges associated with producing plasmid DNA of gene therapy grade is the removal of contaminants such as bacterial chromosomal DNA, RNA, host proteins, and endotoxin.
CHT ceramic hydroxyapatite (Ca5(PO4)3OH)2 support is a leading purification media for today's demanding downstream process industry. Type I CHT media provides high protein binding capacity, especially for acidic proteins.
CHT ceramic hydroxyapatite (Ca5(PO4)3OH)2 support is a leading purification medium for today's demanding downstream process industry. Type II CHT media has lower protein binding capacity but offers better resolution of nucleic acids and is particularly well-suited for the purification of immunoglobulins.
Bio-Gel HT hydroxyapatite is a form of calcium phosphate that is widely used in preparative biochemistry for polyclonal and monoclonal antibody (mAb) separation and the purification of various macromolecules.
Bio-Gel HTP hydroxyapatite is a shelf-stable, dried form of Bio-Gel HT Hydroxyapatite that is ideal for polyclonal and monoclonal antibody separation and the purification of other macromolecules.
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