Using π–π stacking interaction between magnetic Fe₃O₄ graphene oxide nanoparticles and impurities (e.g., biomolecules, free prostate specific antigen (PSA) aptamer-conjugated TEX615 that does not bind with PSA) in human serum, a rapid aptasensor with 1,1′-oxalyldiimidazole chemiluminescence (ODI-CL) detection was developed for the quantification of PSA, a tumor marker. The complexes, PSA bound with PSA aptamer-conjugated TEX615, were formed in human serum while the mixture (1 : 1 volume ratio) of PSA in 25% human serum diluted in Tris–HCl buffer (pH 7.5) and PSA aptamer-conjugated TEX615 in Tris–HCl buffer was incubated for 30 minutes. After the incubation, the impurities capable of reducing the efficiency of the aptasensor were removed using magnetic Fe₃O₄ graphene oxide nanoparticles. With the addition of ODI CL reagents (e.g., ODI, H₂O₂), finally, PSA in each sample was quantified within 1 second. The sensitivity of the aptasensor with good accuracy, precision, and recovery was as good as those of enzyme immunoassays with absorbance and chemiluminescence detections operated using time-consuming procedures such as multiple-time incubation and washing and expensive and intractable antibodies.