Phase tagging affinity separation

Many of the methods developed to study protein interactions use the bait/prey model to detect interacting partners (Phizicky and Fields, 1995 Archakov et al., 2003 Piehler, 2005). The bait protein is a purified protein (often recombinant) that is used to lure and capture a putative interacting protein or biomolecule. The bait protein may be immobilized to a solid phase for affinity separations or be used in solution. It also may be fusion tagged (i.e., GST or 6X His) or labeled with a detectable molecule, such as a fluorescent probe. It often is the case... 

Fluorous affinity separation was originally used to remove catalysts from complex reaction mixtures [6], A perfluoroalkyl moiety (generally no shorter than -C6F13) is appended to a compound of interest. Tagged molecules are then rapidly separated from other components in the mixture by either liquid-liquid extraction or liquid-solid-phase extraction. Fluorous affinity-based separation has recently been used in biomolecule purification, proteomics, and microarray experiments [15-20],... 

A new tag strategy, termed syndiesis based on affinity separation (SAS) , was developed for high throiq)ut synthesis of organic conq)oimds. In this method, the desired tagged conq)ound was separated from the reaction mixture by solid-phase extraction using specific molecular recognition. The interaction between a crown ether (32-crown-lO) and ammonium ion and the interaction between a barbituric acid derivative and its artificial receptor were used for SAS. 

This mode of separation, as the name suggests, uses stationary phases with a special affinity for a specific analyte. The affinity ligand immobilized on the stationary phase varies dramatically from peptide, to protein, to oligonucleotide, to monoclonal antibody. In some cases the target molecule is labelled with an affinity tag to simplify the separation. This approach is common in the synthesis of recombinant proteins where the system can be engineered so that the target biomolecule expresses a tag such as polyhistidine. A stationary phase functionalized with aminodiacetic acid and nickel chelate is then used to fish out the required molecule by chelating with the polyhistidine tag. 

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