The chapter examines the principles of biosensor technology that underpin its application in advanced diagnostics, such as those used for prostate cancer. It presents a summary of what a biosensor is and its central role in transducing a biological response into a measurable signal. The general scheme is as follows: the biorecognition component (bioreceptor) is involved in selective recognition of the analyte by the biomolecule, and the transducer converts this biomolecular process into a measurable physicochemical signal (e.g., electric and optical). The interpretation of the signal handling unit’s results is also elaborated. The chapter presents the sensing mechanism, beginning with the analyte of interest and progressing up to signal recognition. It also ventured into the broad spectrum of biosensors, categorizing them according to the most important top-level classifications, such as the type of bioreceptor (e.g., enzymes, antibodies, and nucleic acids) or transduction principle (e.g., electrochemical, optical, and piezoelectric). The most important performance parameters necessary to test the efficacy and usability of biosensors are presented: sensitivity, specificity, limit of detection (LOD), selectivity, response time, stability, and dynamic range. Ultimately, this chapter effectively demonstrates the significance of these fundamental principles in their real-world applications, which is essential for developing effective biosensors that can accurately identify and detect prostate cancer biomarkers in a timely manner. It serves as a foundation for the more sophisticated designs of biosensors, which are introduced in the second half of the book.