Biodetection using organic field-effect transistors (OFETs) is gaining increasing interest for applications as diverse as food security, environmental monitoring, and medical diagnostics. However, there still lacks a comprehensive, empirical study on the fundamental limits of OFET sensors. In this paper, we present a thorough study of the various parameters affecting biosensing using an OFET decorated with gold nanoparticle (AuNP) binding sites. These parameters include the spacing between receptors, pH of the buffer, and ionic strength of the buffer. To this end, we employed the thrombin protein and its corresponding DNA binding aptamer to form our model detection system. We demonstrate a detection limit of 100 pM for this protein with high selectivity over other proteases in situ. We describe herein a feasible approach for protein detection with OFETs and a thorough investigation of parameters governing biodetection events using OFETs. Our obtained results should provide important guidelines to tailor the sensor's dynamic range to suit other desired OFET-based biodetection applications.