Immunoperoxidase (IP) staining of fine needle aspiration specimens may provide useful adjunct information in cytodiagnosis. IP may be used on unstained direct smears or cell block material if available. In the absence of such material, IP is sometimes used on previously stained (Papanicolaou) smears. Few studies have examined the adequacy of such preparations, however. We compared IP staining results on unstained, alcohol-fixed or air-dried smears versus alcohol-fixed, previously Papanicolaou stained smears and versus alcohol-fixed, decolorized, Papanicolaou-stained smears, using antibodies to keratin (CAM 5.2, AE1/AE3 and K903), epithelial membrane antigen, monoclonal carcinoembryonic antigen, desmin, muscle-specific actin (HHF-35), vimentin, leukocyte common antigen, B and T cell markers L26 and UCHL-1, and prostate-specific antigen. Seventy-five sets of smears were made from various neoplastic or nonneoplastic tissues received as surgical specimens, and antibodies were selected for anticipated positive staining. The results were graded in a semiquantitative fashion. All the methods gave comparable results regarding expected positive staining, background staining and preservation of cellularity. We conclude that these methods of smear preparation are of equal utility in adjunctive immunocytochemical studies.