Label-free immunosensor based on tetra armed star-shaped poly(glycidylmethacrylate) (Star(PGMA)) modified disposable ITO electrode was developed for detection of p53 protein, an important colorectal cancer biomarker. This disposable biosensor was fabricated by spin-coating technique using star-shaped Star(PGMA) with epoxy side groups. After formation of a stable film with epoxy ends, anti-p53 antibodies were bound to these groups covalently. Stepwise modification of the electrode was followed by electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) studies. The electrochemical performance of the immunosensor was studied by EIS. Furthermore, the antibody and antigen coupling was monitored by single frequency impedance (SFI) technique. The immunosensor showed a low limit of detection (7 fg/mL) and a linear detection range between 0.02 pg/mL and 4 pg/mL. Additionally, atomic force microscopy (AFM) and scanning electron microscopy (SEM) were utilized for monitoring of immunosensor surface at different stages of fabrication. The antibody coupling on the electrode surface was proved by Fourier-transform infrared spectroscopy (FTIR) and Raman spectroscopy. Furthermore, the proposed immunosensor had good reproducibility and repeatability.