An electrochemical immunosensor for adiponectin (APN) using screen printed carbon electrodes (SPCEs) modified with functionalized double-walled carbon nanotubes (DWCNTs) as platforms for immobilization of the specific antibodies is reported. DWCNTs were functionalized by treatment with 4-aminobenzoic acid (HOOC-Phe) in the presence of isoamylnitrite resulting in the formation of 4-carboxyphenyl-DWCNTs. The oriented binding of specific antibodies toward adiponectin was accomplished by using the metallic-complex chelating polymer Mix&Go™. The HOOC-Phe-DWCNTs-modified SPCEs were characterized by cyclic voltammetry and compared with HOOC-Phe-SWCNTs/SPCE. The different variables affecting the performance of the developed immunosensor were optimized. Under the selected conditions, a calibration plot for APN was constructed showing a range of linearity extending between 0.05 and 10.0 μg/mL which is adequate for the determination of the cytokine in real samples. A detection limit of 14.5 ng/mL was achieved. The so prepared immunosensor exhibited a good reproducibility for the APN measurements, excellent storage stability and selectivity, and a much shorter assay time than the available ELISA kits. The usefulness of the immunosensor for the analysis of real samples was demonstrated by analyzing human serum from female or male healthy patients.