Novel immunomagnetic microspheres have been prepared for separation of CD4 + lymphocytes. The magnetic nanoparticles with a diameter of approximately 7-8 nm were first synthesized by co-precipitation from ferrous and ferric iron solutions and subsequently entrapped within poly (glycidyl methacrylate) (PGMA) microspheres by a swelling and penetration process. Monoclonal antibody specific to CD4 molecules expressed on CD4 + lymphocytes was conjugated to the surface of magnetic PGMA microspheres through covalent bonding between epoxide functional groups on the microspheres surface and primary amine groups of the antibodies. Flow cytometer was used to confirm the presence of antibody on the surface of microspheres. The new immunomagnetic microspheres are promising to be used in a various biomedical applications including diagnosis, and monitoring of human diseases.