We explored the feasibility of magnetite (Fe 3 O 4 )-gold (Au) core–shell nanocrystals as a useful vehicle for biomedical application such as cell separation. Streptavidin-fluorescein isothiocyanate (STA-FITC) was conjugated to the surface of the Fe 3 O 4 -Au core–shell nanocrystals using a carbodimide activation protocol. These nanocrystals were further tested for their ability to bind CD4 + T lymphocytes, bound to biotin-labeled anti-CD4 mAbs, isolated from the spleen of C57BL/6 mice. Our data show that the Fe 3 O 4 -Au nanocrystals successfully pulled down CD4 + T lymphocytes from the whole splenocytes with high specificity. Therefore, our nanocrystals provide an efficient tool for the cell separation process and further present the dramatic potential to be applied to other areas of biomedical application including diagnosis, monitoring, and treatment of human diseases.