A flexible and reusable SERS substrates were prepared by electrospinning Ag nanoparticles (AgNPs) in reversed micelle into poly(3-caprolactone) (PCL) nanofibers, forming nanocomposite membrane with high detection sensitivity. The AgNPs/PCL nanofiber membrane was characterized by scanning electron microscopy (SEM), transmission electron microscopy (TEM) and surface-enhanced Raman scattering spectroscopy (SERS). SEM and TEM image show that AgNPs dispersed homogeneously in PCL nanofibers. The fabricated substrates based on AgNPs/PCL nanofiber membrane have shown reusability and excellent detection reproducibility in SERS under a low concentration of R6G (10(-12) M) and crystal violet (10(-11) M), and the detection limit of melamine is as low as 5 ppb, which is far below the safety of FDA and US. Moreover, the substrate showed hydrophobicity and low loss of stress after etching.