Bio-Rad offers several protein A media for different mAb purification needs. UNOsphere SUPrA™ Resin is designed for process-scale purification workflows. Affi-Gel® and Affi-Prep® Protein A Resins yield highly purified IgG.
* Refer to Bulletin 3193 for purification conditions. ** For example, Profinity GST Resin is available only in prepacked cartridges. ***Profinity eXact Purification Resin.
Monoclonal Antibody Purification Using UNOsphere SUPrA Media
The first step in purification of an important class of therapeutic proteins, the polyclonal or monoclonal antibodies (mAbs), is their capture from plasma or tissue culture supernatants. Protein A-based media are by far the most common class of affinity products used for this purpose. They bind with high affinity to the Fc region of most subclasses of antibodies and are one of the standard tools used in antibody capture and purification. In combination with ion exchange and ceramic hydroxypatite chromatography, protein A-based media have been successfully used in the large-scale purification of numerous licensed mAb drugs.
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* 10% breakthrough capacity determined with 1.0 mg/ml polyclonal human IgG in 1.1 x 10 cm column.**No significant change in chromatographic performance
* Approximately 50% of all mouse IgMs bind using the MAPS buffer system.
The Profinia Protein Purification System Simplifies Antibody Purification with Protein A
Protein A affinity purification (–) and desalting (–) profiles using 0.1 M citrate elution buffer.
Comparison of protein A elution profiles using 0.1 M glycine, pH 3.0 (–) and 0.1 M citrate, pH 3.0 (–) elution buffers.
Protein A purification profiles using three concentrations of glycine, pH 3.0 (0.1 M (–); 0.25 M (–); 0.5 M (–)) for elution.
Protein A affinity purification (–) profiles using 0.25 M glycine elution buffer.
Purification of IgG from human serum using the Profinia protein purification system. Left panels represent chromatograms from purifications of IgG using 1 ml of human serum. The Profinia system monitors the absorbance at 280 nm from the affinity and desalting cartridges. Panels on the right display images from SDS-PAGE analyses of protein fractions from each experiment. M, Precision Plus Protein standards; L, load; F, flowthrough; W, wash; E, elution. The gel in D includes IgG purified using 0.1 M citrate as the elution buffer for comparison.
A Method for Rapid, Large-Scale Removal of Albumin and IgG from Human Serum Using the BioLogic DuoFlow Chromatography System
In human serum, albumin contributes more than 60% of total protein, and immunoglobulins, predominantly IgG, contribute 10–25%. The high abundance serum proteins and limit the amount of total serum protein that can be resolved by two dimensional (2-D) electrophoresis.
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