Magnetic FeNi3 nanochains were synthesized by reducing iron(III) acetylacetonate and nickel(II) acetylacetonate with hydrazine in ethylene glycol solution without any template under a rapid and economical microwave irradiation. The morphology and composition of the as-prepared products were characterized by field-emission scanning electron microscopy (FESEM), transmission electron microscopy (TEM), X-ray diffraction (XRD), and elemental mapping. The size of the aligned nanospheres in the magnetic FeNi3 chains could be adjusted from 150 to 550 nm by increasing the amounts of the precursors. The length of the nanochain is about several tens of micrometers. The ratio of the precursors plays an important role in the formation of FeNi3 nanostructures. Magnetic measurement reveals that the FeNi3 nanochains show enhanced coercivity and saturation magnetization. The formation mechanism of the product is discussed. Toxicity tests of FeNi3 nanochains show that the as-prepared nanochains are nontoxic to zebrafish larvae. In vitro magnetic resonance imaging (MRI) confirms the effectiveness of the FeNi3 nanochains as sensitive MRI probes.