Carbon nanofibers, with diameters between 80 and 290 nm and specific area of 242 m2g-1, have been prepared by the catalytic chemical vapor deposition method. After preparation, the powder was mixed with silicon oil in order to create a paste electrode. The electrochemical behavior of this type of electrode was investigated by cyclic voltammetry, using a solution of 10-3 M ferrocenecarboxylic acid as mediator. The redox process is quasi-reversible, and it involves the transfer of electrons between Fe(II) and Fe(III). The same mediator was used for the construction of a second-generation glucose biosensor. The mediator was co-immobilized with the enzyme in the carbon nanofibers paste. The sensor linearly responded to glucose, in the concentration range of 1.7 to 7 mM. A time of around 30 seconds was required to reach 95% of the maximum steady-state current. Also, the oxidation of calf thymus DNA at the carbon nanofiber paste electrode was investigated by differential pulse voltammetry (DPV). A clear signal, due to guanine oxidation, was obtained in the case of single-stranded DNA.