This work evaluates the effect of purification process and storage conditions (buffer formulation and temperature) on the efficacy of Ehrlichia ruminantium (ER) elementary bodies to be used as an inactivated vaccine candidate against heartwater. In vitro assays revealed that, to avoid major losses in ER integrity and corresponding antigenic properties, a buffer with pH between 5.6 and 8 and an osmolality above 100 mOsmol/kg H(2)O is recommended. Amongst the tested formulations, both PBS and NaCl have shown to stabilize ER antigens at -20 degrees C. To assess the protective properties of the different vaccine formulations, in vivo experiments were performed using a goat model. The results obtained showed that the preparation of ER antigens using a novel membrane-based purification strategy and a simple vaccine formulation (NaCl, -20 degrees C) induced equivalent protection to the conventional vaccine based on ER antigens prepared by a multistep centrifugation methodology and stored at -20 degrees C in PBS buffer.