We describe a method of immunocytochemically assessing estrogen receptor (ER) status on alcohol-fixed smears obtained by fine-needle aspiration (FNA) from breast cancer patients, using a commercially available monoclonal antibody (1D5) with microwave oven processing. A series of 31 cases of aspirates from breast cancer were analysed and the results were compared with assessment by ER immunocytochemical assay using the same procedure on formalin-fixed tissue and with assessment by ER-ICA assay on frozen sections. The results were scored semiquantitatively using a five grade scoring system. Of the 31 cases examined, 21 were positive at least by two methods and 10 were negative for all three determinations. The results obtained in the ER immunocytochemical assay on aspirates and paraffin-sections using the antibody 1D5 and those obtained on frozen sections using the antibody H222 were closely similar. In only one case was it not possible to interpret the reaction in the cytological specimen because there was a strong background in the smear. In general, we obtained more intense positivity with the antibody 1D5 in aspirates and formalin-fixed material than with the antibody H222 in frozen sections. The scoring results of the three methods were almost identical. We conclude that the application of ER method on alcohol-fixed smears will eliminate the need for using a special fixation procedure and will provide several advantages, such as: improvement in morphological concomitant analysis, utilization whenever malignancy is found without necessity to re-aspirate the patient, and adequacy of archival material. © 1995 Wiley-Liss, Inc.