The electrochemical behavior of cytochrome c on gold electrodes modified in solution containing a mixture of 3-mercaptopropionic acid and n-alkanethiol having various chain lengths was investigated and compared with that of another mixed monolayer system, bis(4-pyridyl) disulfide and n-alkanethiol, previously reported. The peak current decreased with the increase of the ratio of n-alkanethiol in the 3-mercaptopropionic acid + n-alkanethiol mixture, as was observed in the case of bis(4-pyridyl) disulfide + n-alkanethiol mixture. The n-alkanethiol with a longer chain was more effective in blocking the electron exchange between cytochrome c and the electrode than that with a shorter chain, in contrast to the results observed at the mixed monolayer of bis(4-pyridyl) disulfide and n-alkanethiol. This observation reflects the fact that adsorbed molecules were well mixed with each other in the case of the 3-mercaptopropionic acid + n-alkanethiol mixed monolayers, while patched domains of pyridyl thiolate were formed in the case of the mixed monolayers of bis(4-pyridyl) disulfide and n-octadecyl mercaptan. The electrochemical behavior of the gold electrodes modified with the mixed monolayer of 3-mercaptopropionic acid and n-alkanethiol in the solution containing Fe(CN)(6)(4-/3-) support the above observation. Reductive desorption of the mixed monolayer was carried out in alkaline solution to determine the composition and to evaluate the degree of mixing of the two components.