AP188P Sigma-AldrichMouse Anti-Rabbit IgG Antibody, HRP conjugate, Species Adsorbed

We report a simple method to microencapsulate erythropoietin (EPO, a protein easily denatured and antigenized by contact with water-organic solvent interfaces) into poly(lactic-co-glycolic acid) (PLGA) microspheres with minimal aggregation. This formulation process involved an aqueous-aqueous emulsion formed at reduced temperature. EPO was first dissolved in water together with dextran (MW=70,000) and polyethylene glycol (MW=8000), followed by a freezing process during which dextran separated out as the dispersed phase with EPO partitioned in preferentially. The frozen sample was then lyophilized to powder and washed with dichloromethane to remove the PEG continuous phase. Once loaded in the polysaccharide particles, 1-4 microm in diameter, EPO gained resistance to organic solvents and was encapsulated into PLGA microspheres without significant aggregation (2%). EPO released from the composite PLGA microspheres in a sustained-release manner with minimal burst (20% at the first day) and incomplete (20%) release. Single injection of these EPO-loaded PLGA microspheres to mice resulted in a red blood cell elevation equivalent to twelve injections of the solution formulation. An ELISA assay suggested that the mice injected with the EPO-loaded PLGA microspheres did not develop anti-EPO IgG more than those given solution form EPO.