Electrochemical biosensor for salivary RNA detection with high sensitivity is presented. The complexity of saliva and the low concentration of salivary RNA require both high sensitivity and specificity. In this study enzyme amplification is applied to achieve high signal intensity. In order to increase the current signal, effective hybridization and electron transfer are both required. Electrochemical studies on the location of anchor label and report label have been carried out. Results indicate the hybridization efficiency and the surface hindrance to enzyme amplification are the two major factors contributed to the detected current signal. Human salivary IL-8 RNA has been detected. For oligonucleotide target, the limitation of detection (LOD) is about 1 fM, while the LOD for IVT RNA is about 50 pM.