One of the most important problems faced by microbiologists is to preserve bacterial isolates in the best state to study and further diagnosis. The current study aims to provide a summary of experimental results to maintain two species of bacteria alive after being stored by using some additives. This study found that the best temperature to preserve Staphylococcus aureus was -20°C for a year, while for Escherichia coli it was the same temperature except in using Glycerol (G) 100% and Food oil (FO) methods. The optimum method to preserve S. aureus was by using Normal Saline (NS), while Distilled Water (DW) was the optimum method to preserve E. coli at temperatures (4, 25 and -20)°C for a year, the phenotypic patterns for examining bacteria were maintained except in NS at 4°C for S. aureus after a year ago. Glycerol was used alone at concentrations (100, 50, 30 and 15)%, and another group used G+NS in the same volumes, good results were achieved when it used alone or with NS to preserve bacteria for six months at 4°C except for methods of G100% and (G100% + NS) for examining bacteria. FO has never been used as preservation liquid, it is successful to survive S. aureus at -20°C for a year, and when it was added to NS, E. coli survived for a year at three temperatures (4, 25 and -20)°C, while S. aureus didn’t survive for a year when FO+NS method used at room temperature. The precipitation method was used for bacterial suspension, then added the preserving liquid, but the results were not effective compared to the First method.