Magnetic nanoclusters (MNCs) are developed and used for an ultrasensitive magnetophoretic assay system. Prostate‐specific antigen (PSA) is used as a target analyte (see image). The deflection velocity of a microbead is well correlated with the PSA concentration, with a detection limit of 45 fg mL−1, and this value is three orders of magnitude higher than conventional ELISA. The developed assay format is expected to be effectively applied to the detection of a target analyte with low abundance.