Imaging of gold colloidal particles bound to immune complexes on conductive substrates was carried out with scanning tunneling microscope (STM) operated in air. STM images of the gold colloids were obtained on models of immunoassay sample of ‘‘gold colloidal particle/immune complex/substrate.’’ One model is composed of gold colloidal particles (average diameter of 15 nm) and an antigen–antibody complex [human immunoglobulin G(IgG) and rabbit anti‐human IgG] on a gold substrate. The other the same kind of gold colloids and another kind of immune complex [Hepatitis B surface (HBs) antigen and anti‐HBs]on a highly oriented pyrolytic graphite. It was possible to obtain the STM images of gold colloids bound to the immune complexes without heavy metal coatings. The STM images showed corrugations consistent with the coverages expected from the nominal size of the gold colloidals used. The successful measurement suggests that we can make use of STM for a novel immunoassay, ‘‘STM immunoassay,’’ for detecting a trace amount of macromolecules of biological importance.