In this study, we have developed a novel paper based immunoassay for the quantitative detection of immunoreactions using electrochemical impedance spectroscopy. Paper provides an attractive platform for fabrication of simple, low cost, and portable diagnostic devices as it allows passive liquid transport, is biocompatible, and has tunable properties such as hydrophilicity, flexibility, permeability, and reactivity. We have used screen-printing to fabricate interdigitated electrodes (finger width and gap of 200 μm) on the paper substrate, while UV-lithography enables patterning of the paper into hydrophobic/hydrophilic regions. As a proof of concept, we have used this immunosensor to detect the immune response of Human Serum Albumin (HSA) antibody-antigen complex formation. To enable efficient immobilization of HSA antibodies, we have utilized dielectrophoresis to trap microprobes (MPs) on the electrode surface. The microprobes consist of an alumina nanoparticle core with a well-adhered polyaniline outer coating to which the HSA antibodies are conjugated in an oriented manner via covalent chemistry. The efficacy of the impedance-based immunosensor is compared when MPs are immobilized specifically on the electrode surface using dielectrophoresis (DEP) as opposed to being dropped and immobilized via physical absorption on the entire sensing area. Results show that a more reproducible and sensitive response is observed when DEP is utilized to trap the microprobes. Furthermore, the normalized impedance variation during immunosensing shows a linear dependence on the concentration of HSA with an observed limit of detection of 50 μg/ml, which is lower than conventionally used paper based urine dipsticks used for urinary protein detection. Thus, we have developed a low cost paper based immunoassay platform that can be used for the quantitative point of care detection of a wide range of immunoreactions.