With the help of iron oxide nanoparticles, electron spin resonance spectroscopy (ESR) was applied to immunoassay. Iron oxide
nanoparticles were used as the ESR probe in order to achieve an amplification of the signal resulting from the large amount of
Fe3+ ion enclosed in each nanoparticle. Rabbit IgG was used as antigen to test thismethod. Polyclonal antibody of rabbit IgG was
used as antibody to detect the antigen. Iron oxide nanoparticle with a diameter of either 10 or 30 nm was labeled to the antibody,
and Fe3+ in the nanoparticle was probed for ESR signal. The sepharose beads were used as solid phase to which rabbit IgG was
conjugated. The nanoparticle-labeled antibody was first added in the sample containing antigen, and the antigen-conjugated
sepharose beads were then added into the sample. The nanoparticle-labeled antibody bound to the antigen on sepharose beads
was separated from the sample by centrifugation and measured.We found that the detection ranges of the antigen obtained with
nanoparticles of different sizes were different because the amount of antibody on nanoparticles of 10 nm was about one order of
magnitude higher than that on nanoparticles of 30 nm. When 10 nm nanoparticle was used as probe, the upper limit of detection
was 40.00 μg mL−1, and the analytical sensitivity was 1.81 μg mL−1. When 30 nm nanoparticle was used, the upper limit of
detection was 3.00 μg mL−1, and the sensitivity was 0.014 and 0.13 μg mL−1 depending on the ratio of nanoparticle to antibody.

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• Electron spin resonance spectroscopy for immunoassay using iron oxide nanoparticles as probe