By PHARMACIA FINE CHEMICALS
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Extra resources for Affinity Chromatography: Principles and Methods
Protein G also binds rat IgG2a and IgG2b, which either do not bind or bind weakly to protein A. Amersham Biosciences offers a recombinant form of protein G from which the albuminbinding region of the native molecule has been deleted genetically, thereby avoiding undesirable reactions with albumin. Recombinant protein G contains two Fc binding regions. Protein G Sepharose is a better choice for general purpose capture of antibodies since it binds a broader range of IgG from eukaryotic species and binds more classes of IgG.
Elute the IgY with 10 column volumes of elution buffer. 6. Wash the column with 8 column volumes of wash buffer. 7. Immediately re-equilibrate the column with 5 column volumes of binding buffer. 8 M Na2SO4. The sample should have the same concentration of Na2SO4 as the binding buffer. An increase in salt concentration will reduce the purity of the eluted IgY. The purity of the eluted IgY may be improved by using gradient elution with, for example, a linear gradient 0–100% elution buffer over 10 column volumes, followed by elution with 100% elution buffer for a few column volumes.
Supplied as a suspension ready for column packing. 500 cm/h** For fast processing of large sample volumes. Retains high binding capacity at high flow rates. Supplied as a suspension ready for column packing. MabSelect™ Human IgG, approx. 30 mg/ml medium (recombinant protein A ligand) *Protein A Sepharose 4 Fast Flow and rProtein A Sepharose Fast Flow have a higher binding capacity, a more rigid matrix and provide more convenient alternatives to Protein A Sepharose CL-4B, which must be rehydrated before column packing.
Affinity Chromatography: Principles and Methods by PHARMACIA FINE CHEMICALS