A self-assembled highly oriented P3HT (Poly(3-hexylthiophene-2,5-diyl))/GO (Graphene oxide) nanocomposite-based p-channel organic field-effect transistor (OFET) for real-time ammonia sensing has been fabricated and characterized on a p++ (highly boron-doped) silicon substrate at room temperature (∼25 °C). A low-cost facile floating film transfer method has been adopted to fabricate the sensing channel of OFET with optimized and uniform thickness (∼25 nm). The electrical characterizations and NH3 sensing performance of the P3HT/GO nanocomposite OFET with palladium electrode have been characterized and compared with pristine P3HT-based OFET. It is demonstrated that the polymer nanocomposite-based OFET with Pd electrode has a relatively high response of ∼63% at 80 ppm NH3 gas. A reduction of 60.41% in field-effect mobility (0.0551 cm2/ V. sec. to 0.02181 cm2/V. sec.) and a 29.4% change in threshold voltage (−4.758 V to −6.157 V) has been observed by varying the NH3 gas concentration over 0–80 ppm. It has an improved response time of ∼44 s and recovery time of ∼82 s due to improved crystallinity and grain size of the sensing layer. All the electrical characterizations and sensing measurements have been done at RT and 55% relative humidity.