Boronate affinity chromatography of cells and biomacromolecules using cryogel matrices

Abstract

Boronate affinity chromatography involves the interaction between cis-diol containing molecules and the hydroxyl group of boronate. Boronate affinity based cryogel chromatography matrices have been developed and the ligands were immobilized by two methods i.e., grafting of the boronate ligand on to the matrix and by copolymerization of monomer containing boronate with other co-monomer. The boronate grafted cryogel column was used to capture adherent and non-adherent cells and the captured cells were recovered at different fructose concentrations as an eluting agent, in chromatography mode. It was found that the adherent cells could be recovered at relatively higher fructose concentration (0.5M) than non-adherent cells which could be recovered by using low fructose concentration (0.1M). This might be due to the difference in the content of glycoprotein in adherent and non-adherent cells. In this way a new separation method can be devised for the fractionation of adherent and non-adherent cells. In another study, a copolymerized boronate cryogel column was developed for the separation of RNA from the bacterial crude extract without any pre-processing. RNA molecules were specifically retained in the cryogel column due to interaction between 2,3′ diol group of ribose sugar in RNA and the hydroxyl group of boronate. The DNA molecules were passed through the column uninteracted due to absence of 2′-hydroxyl group. Later, bound RNA molecules were recovered from the boronate affinity cryogel column.