Electrochemical biosensors are portable devices that permit rapid analysis of substances. They are most useful in detection and monitoring of biological, chemical and toxic agents. Briefly, with the help of transducer, the generated electrical signals from the responses to change in the bioactive layers are used for the interpretation. Similarly, nanomaterials have number of features that make them ideally suited for sensor applications, such as, its high surface area, high reactivity, easy dispersability and rapid fabrication. This collected work composed of the expert knowledge of many specialists in the construction and use of electrochemical biosensors made of nanostructured materials. This includes nanomaterials such as dendrimers, polymers, nanoparticles, nanotubes, oxides, enzymes and their hybrids as catalyst for various sensors such as glucose sensors, DNA sensors, neurotransmitters sensors, etc. This collected work provides new methodological advancements related to and correlated with the measurement of interested species in biomedical samples. Many studies are also included to illustrate the range of application and importance of the electrochemical biosensors. This provides the unique opportunity for readers to choice a new methods and applications of new electrochemical biosensors.