Sensitive, accurate, and rapid detection of biomolecules from complex matrices still remains a critical challenge. Herein, we designed magnetic nanoparticles which were coupled with plasmonic surfaces to detect a clinically important biomarker (interferon-alpha) in buffer solutions and artificial plasma samples. At first glance, the magnetic nanoparticles were synthesized and comprehensively characterized. Then, any potential external factors on magnetic nanoparticles were examined, thereby benchmarking detection performance. After determining optimum conditions, the plasmonic biosensor surface was modified with these magnetic nanoparticles for detecting interferon-alpha in real-time. The kinetic performance of magnetic nanoparticles-assisted plasmonic biosensor provided a high correlation coefficient (0.99) and a low limit of detection (41 IU/mL) in a broad range of interferon alpha concentrations. Consequently, this method holds a great potential to detect other biomolecules with high sensitivity and low interference and would be employed as an alternative approach for biomarker detection at different perspectives ranging from medical diagnosis to pharmaceutical industry.