Investigating protein-protein interactions (PPIs) is essential in basic and clinical research, but current techniques for detecting PPI are hampered by lengthy protocols, reduced sensitivity, and a high rate of false positive results. To overcome these challenges, we use a magnetic modulation biosensing (MMB) system, in which one protein of interest is attached to a magnetic bead and another protein is coupled to a fluorescent molecule. Thus, when the two proteins interact, the fluorescent molecule is connected to the magnetic bead. An oscillating magnetic field gradient then concentrates the beads and transports them in a periodic motion in and out of a laser beam, producing an oscillating signal that is detected and demodulated. Using recombinant and native proteins, we identified the known interaction between erythropoietin and its receptor. In addition, we demonstrated for the first time the direct interaction of a recently identified and characterized toxin antitoxin pair in Pseudomonas aeruginosa. Compared to the commonly used Co-immunoprecipitation technique coupled to western blot, which takes approximately 10 h, a measurement in the MMB system takes only two minutes and has much less non-specific binding. We anticipate that the MMB-based technique will significantly advance the detection of PPIs in medical and drug discovery research.