We reported the controlled surface morphologies and the cell culture of polyelectrolyte multilayer coated nylon 6 fibrous mats with different number of layers. Polyelectrolyte multilayer coated nylon 6 fibers were successfully prepared by an alternative deposition of alginic acid sodium salt and chitosan via a Layer-by-Layer (LbL) electrostatic self-assembly. The surface morphology, stiffness, and hydrophilicity of polyelectrolyte multilayer coated nylon 6 fibrous mats could be finely tuned by regulating the number of polyelectrolyte nanocoating. It was observed that the morphology of polyelectrolyte multilayer coated nylon 6 fibers was uniform and smooth, indicating a dense and harder nanocoating of polyelectrolytes onto nylon 6 fibers. Compared to pure nylon 6 fibrous mat (tensile strength ∼10.6±1 MPa), the tensile strength of polyelectrolyte coated nylon 6 fibrous mats was largely increased to 35.2±2 MPa for 5 bilayers coated fiber mats. In addition, it was found that at an initial stage after 1 day of cell culturing, the electrospun nylon 6 fibrous mats coated with 5 bilayer of alginic acid and chitosan show the highest cell affinity (good adhesion), while the electrospun nylon 6 fibrous mats coated with 10 bilayer show the lowest cell affinity. After cell seeding for 3 days, it was observed that rate of proliferation is enhanced by increasing the number of bilayer up to 3 bilayers (good proliferation), and then drastically decreased with further increasing the number of bilayer.