Zika virus (ZIKV) recently emerged as a global threat subsequent to its global spread because it induces microencephaly and other brain damages in infants born to infected mothers. Epidemiological monitoring of infection has been hampered by the absence of reliable serological tests capable to distinguish between ZIKV and other Flavivirus infections， in particular Dengue virus (DENV). As both viruses are transmitted by the same mosquito-species， their distributions largely overlap and reliable serological distinction between the viruses is essential. Here we develop a novel biosensor which is based on recombinant forms of ZIKV non-structural protein 1 (NS1) and the domain III of the envelope protein (EDIII). Using electrochemical impedance spectroscopy (EIS) and square wave voltammetry (SWV)， we demonstrate that in addition to extremely sensitive detection of ZIKV-specific antibodies in serum and saliva， the biosensor promptly distinguished ZIKV and DENV-specific antibodies. Hence， this novel biosensor allows assessing ZIKV antibodies in blood and saliva and results are unaffected by presence of DENV virus-specific antibodies.