In this work, a novel hydrothermal-calcination process was introduced for the fabrication of sheet-like α-Fe
2
O
3
/Fe
3
O
4
magnetic nanocomposites with an average diameter of 220 nm, average thickness of 130 nm, and saturation magnetization of 27.6 emu∙g
-1
. With the nanosheets as the precursors, the α-Fe
2
O
3
/Fe
3
O
4
@Au-ssDNA/BSA probes with label-free and magnetically induced self-assembly were constructed for the detection of circulating tumor DNA. The probes revealed excellent linear relationship as I = -2.62 lgC
ctDNA
+138.35 (the linear range of 10 pM–1 μM for the concentration of circulating tumor DNA) with an R
2
of 0.998, lower limit of detection of 229 fM, and limit of quantification of 763 fM. The biosensors revealed superior reproducibility (relative standard deviation of 1.13%), excellent stability (the detection errors of -0.97%–1.47% in 14 days), and high selectivity, suggesting the significant clinical application value in the detection of circulating tumor DNA for the biosensors.