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We have evaluated the use of harmonic signals for detecting magnetic nanoparticles (MNPs) in biomedical applications. We studied the relationship between the harmonic signals and the distribution of magnetic moments in MNP samples. We measured the harmonic signals and frequency dependence of the ac susceptibility along with static M-H curves. These experimental results were analyzed to obtain the magnetic moment distribution in real samples. We were able to clarify what proportion of MNPs contributes to the harmonic signal. MNPs could be classified according to three types. Type-1 contributes to the fundamental signal only. Type-2 contributes to both the fundamental and harmonic signals. Finally, Type-3 does not contribute to any signals. It is important to choose a sample having a large portion of Type-2 MNPs for the harmonic signal-based detection.