Stress biomarkers such as hormones and neurotransmitters in bodily fluids can indicate an individual’s physical and mental state, as well as influence their quality of life and health. Thus, sensitive and rapid detection of stress biomarkers (e.g., cortisol) is important for management of various diseases with harmful symptoms, including post-traumatic stress disorder and depression. Here, we describe rapid and sensitive cortisol detection based on a conducting polymer (CP) nanotube (NT) field-effect transistor (FET) platform. The synthesized polypyrrole (PPy) NT was functionalized with the cortisol antibody immunoglobulin G (IgG) for the sensitive and specific detection of cortisol hormone. The anti-cortisol IgG was covalently attached to a basal plane of PPy NT through an amide bond between the carboxyl group of PPy NT and the amino group of anti-cortisol IgG. The resulting field-effect transistor-type biosensor was utilized to evaluate various cortisol concentrations. Cortisol was sensitively measured to a detection limit of 2.7 × 10−10 M (100 pg/mL), with a dynamic range of 2.7 × 10−10 to 10−7 M; it exhibited rapid responses (<5 s). We believe that our approach can serve as an alternative to time-consuming and labor-intensive health questionnaires; it can also be used for diagnosis of underlying stress-related disorders.