In the past years, number of reports have documented the endocrine disrupting activity of steroid hormones including progesterone, a potent lipophilic compound which is essential for the diagnosis of various reproductive health disorders and hormonal imbalance. Owing to the admirable physio-chemical characteristics, graphene and its functional nanocomposites have garnered a considerable attention in the domains of electrochemical sensing. Herein, the proposed work presented the usage of Magnetic nanocomposites of graphene oxide (MGO) as electrode casting material for constructing immunosensing platform for progesterone hormone. MGO decorated screen printed electrodes were further implemented as bio receptors-immobilization matrix due to its enhanced electrochemical response as well as the availability of plenty of active groups for binding antibodies. The electrochemical performance was investigated with both Cyclic Voltammetry (CV) and Differential Pulse Voltammetry (DPV) techniques for effective detection and analysis. Under optimum experimental parameters, the present method displayed excellent repeatability, selectivity, fine stability and a good linearity from 0.01 pM to 1000 nM with low detection limits i.e., 0.15 pM (with DPV) and 0.17 pM (with CV). Subsequently the developed biosensor was applied in real sample analysis practically with spiked water samples and exhibited satisfactory recovery performance. Furthermore, the structural properties of MGO were studied through various characterization methods such as FE-SEM, FTIR, XRD, and RAMAN. Application of two electrochemical techniques in the study has significantly improved the sensor’s performance as cost effective as well as reliable detection approach in biological matrices.