Peptides with unique features such as small molecular weight, ease synthesis, good biocompatibility, high stability and versatile sequences, have received increasing interest as recognition elements for biosensors. Electrochemical analysis has wide applications because of its high sensitivity, good accuracy, simple equipment, wide detection range and easy to use. The peptide over bar based electrochemical biosensors can be used in many fields including environment monitoring, biomedicine and food detection. In this review, we introduce the peptide over bar based electrochemical biosensors for detection heavy metal ions, small over bar molecules, proteins, pathogenic bacteria and viruses. The strategies for peptide modification and immobilization are summarized. We also describe the properties and sensing mechanisms of peptide over bar based electrochemical biosensors. Sensing strategies especially those based on target over bar binding induced combination, digestion and phosphorylation of peptides have been discussed. Lastly, the current problems and prospect of the research on peptide over bar based electrochemical biosensor are discussed and prospected.