SOL-GEL IMMOBILIZED TIOPHILIC LIGANDS AND PROTEIN A IN THE RECOVERY OF THE GAMMA FRACTION OF HUMAN SERUM

M.V. TUTTOLOMONDO; M.E. VILLANUEVA; DIAZ, L

Congreso; XLVI Reunión Anual Sociedad Argentina de investigación en Bioquímica y Biología Molecular; 2010

Interest in monoclonal and polyclonal antibodies has increased as more applications in biotechnology such as immunoaffinity chromatography, immunodiagnostic, drug targeting and biosensors are found for these versatile molecules. For all these purposes homogeneous antibody preparations are needed. Conventional purification methods using Protein A are expensive to operate due with. None-specific methods like hydroxyapatite or ion exchange chromatography possess low capacity or yield antibodies contaminated with albumin and transferrin. The use of tiophilic ligands shows selective binding of immunoglobulins in the presence of structure forming salts. The most common sorbent structure derives from divinylsulphone coupled with Beta-mercaptoethanol. For the purpose of comparing both techniques, glass slides and glass beads were functionalized with APTES, which provide ?NH groups 2 available for reaction with glutaraldehyde (needed for Protein A attachment) and divinylsulphone (needed for the coupling with beta- mercaptoethanol). The adsorption of Gamma- fraction from human serum is being evaluated, using capillary electroforesis, as a Gamma- fraction:albumin ratio change. Desorption of the adsorbed proteins and reutilization of the derivatized glass slides is also being evaluated