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Introduction: Brain (central nervous syst-em) is responsible to manage all biological process in living organisms, particularly mammalian. Since proteins play major roles in the biological cell activities, nowadays identification of brain proteins and deter-mination of their function is interesting subject for neurologists. Isoelectric Foc-using (IEF) technique is one of the prote-omic technologies to study brain proteins. This study was carried out to select the IPG strips with maximum resolution and to optimize IEF condition for the separation of mouse brain proteins. Materials & Methods: After extraction of mouse brain proteins and determination of protein concentration by Bradford assay, Isoelectric Focusing was performed using IPG strips with three different pH gradients. Findings: IPG strip with pH=3-10NL was able to separate more proteins with relati-vely high resolution in comparison with IPG strips with pH=3-10 and pH=4-7. Discussion & Conclusion: Results were shown that more proteins can be separated using IPG strip with pH=3-10 NL and this strip has a high resolution rather than the other strips. Due to the complexity of brain, its function, and the extent of brain proteins, it is appropriate to resolve proteins by the method.