Summary: Developing rapid detection tools and methods for foodborne pathogens has always been a research hotspot, which is undoubtedly very important for food safety. In this study, the preparation of functional magnetic nanoprobes and their application to detect target pathogens in a complex matrix by low‐field nuclear magnetic resonance (LF‐NMR) is demonstrated. The long‐arm functional magnetic nanoprobes were constructed by modification with polyethylene glycol (PEG) onto carboxyl‐modified magnetic nanoparticles (MNPs) and then reaction with biotin (Bio). Under the optimal conditions, the lower limit that can be detected in the PBS buffer system is 100 CFU/mL, and the detection of the actual sample matrix can be achieved 102 CFU/g. The practical application of LF‐NMR sensors to detect harmful pathogenic bacteria in food provides an efficient way of monitoring dangerous pathogenic diseases. [ABSTRACT FROM AUTHOR]