Concanavalin A Binding on PHEMA Beads and Their Interactions with Myeloma Cells

Abstract

The aim of this study is to prepare concanavalin A (Con A) bound poly(2-hydroxy ethyl methacrylate) (PHEMA) beads for cell affinity chromatography. In the first step, PHEMA beads were produced by suspension polymerization, and activated by cyanogen bromide (CNBr) in an alkaline medium (pH 11.5), and then, the bio-ligand “Con A” was attached by covalent binding onto the CNBr activated beads. PHEMA beads were characterized by scanning electron microscopy (SEM), surface area and pore size measurements. The PHEMA beads have a spherical shape and porous structure. The specific surface area of the PHEMA beads was found to be 39.7 m2/g with a size range of 150–200 μm in diameter and the swelling ratio was 55%. The amount of bound Con A was controlled by changing pH and the initial concentrations of CNBr and Con A. The non-specific adsorption of Con A on the plain PHEMA beads was 0.1 mg/g. The maximum Con A binding was 4.8 mg/g at pH 7.25. Both plain and Con A bound PHEMA beads were interacted first with the myeloma cell suspension in phosphate buffer. Myeloma cell attachment was very low for the plain PHEMA beads, while the number of myeloma cells attached increased almost 20 fold when the Con A bound beads were used. In order to look at whether or not the interaction of the Con A bound PHEMA beads and myeloma cells are affected from the biological molecules and other cells in the medium. We selected sheep blood itself as the medium, and mixed with the myeloma cell suspension and changed the environment. Cell adhesion decreased but not very significantly by changing the medium from simple buffer to sheep blood.