Stabilization of biomolecules on matrices is critically important. Here we constructed composite nanofibers with magnetic features by electro-spinning of magnetic nanoparticles (MNP) in an appropriate polymer matrix on a collector surface for multiple uses. 'Poly(vinylalcohol)-polyethyleneimine/Fe3O4 magnetic nanoparticles' (PVA-PEI/MNP) composite nanofiber was used on the screen printed electrodes (SPE) to serve as a platform for the glucose oxidase (GOx) biofilm as the model biomolecule. To show the potential application of this material as a biosensor component, the resulted biofilm, called 'PVA-PEI/MNP/GOx', was fixed on SPE via a neodymium magnet for the electrochemical detection of glucose at -0.7 V where oxygen consumption due to enzymatic reaction was measured. The main advantage of the obtained magnetic biomembrane is that it allows analysis with a single drop. After step-by-step surface modifications, analytical characterization was performed using various techniques, such as voltammetry, electrochemical impedance spectroscopy, Fourier-transform infrared spectroscopy and X-Ray photoelectron spectroscopy as well as microscopic techniques. The linear range of the PVA-PEI/MNP/GOx surface was found as 0.0125 to 0.5 mM with a limit of detection of 11.5 mu M for glucose as analyte. The application of PVA-PEI/MNP/GOx for glucose detection in synthetic samples was carried out. Data proved the potential use of the 'PVA-PEI/MNP' surface as a biomolecule immobilization platform for disposable biosensor applications.