Magnetic nanoparticles (NPs) can serve as magnetic relaxation switches (MRSw's), switching from a dispersed to a clustered state, or the reverse, due to the presence of molecular targets, with changes in the spin-spin relaxation time of water (T_2). Biotinylated NP probes reacted with an avidin molecular target to form stable NP clusters, which permitted several NMR parameters to be measured as a function of cluster size. Associated with avidin-induced NP cluster formation was an increase in the spin-spin relation rate (1/T_2), while the spin-lattice relaxation rate (1/T_1)was unaffected. On the basis of the selective effects of NP cluster formation on T_2, we developed a T_1/T_2 interrogation method where NP probe concentration and avidin analyte were unknown and both were determined. A third NMR parameter examined was the replication of T_2 measurements, which were used to rapidly determine whether the ratio of avidin to biotinylated NP was optimal or whether additional biotinylated NP was needed. The T_1/T_2 and T_2 replication interrogation methods illustrate how MRSw assays can employ multiple parameters, instead of relying only on T_2, to obtain information about the reaction of NPs with molecular targets.
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