The value of immunohistochemical staining in the subtyping of acute leukemia was investigated on 36 routinely processed (formalin-fixed and paraffin-embedded) trephine biopsy specimens from the iliac crest containing diffuse infiltrates of acute myelogenous leukemia (AML; n = 23) and acute lymphoblastic leukemia (ALL; n = 13). These were stained with a broad panel of antibodies (n = 23) against various leukocyte antigens, among them 11 macrophage-associated antibodies (MAAs): Ki-M1p, MAC387, HAM56, LN5, KP1 (CD68), PG-M1 (CD68), Ki-M4p, DAKO-DRC (CD35), and antibodies against lysozyme, alpha 1-antichymotrypsin, and S100 protein. The French-American-British (FAB) classification subtypes of the AML cases, as determined by enzyme-cytochemical and/or immunocytological investigation of bone marrow smears, were as follows: M1 = 6, M2 = 5, M4 = 7, M5 = 3, and AML (not classified) = 2. The 13 cases of ALL were classified as follows: c-ALL (pre-B-ALL) = 7, B-ALL = 3, T-ALL = 2, and ALL (not classified) = 1. All the MAAs except LN5, Ki-M4p, and DAKO-DRC stained blast cells in AML. However, the number of stained blast cells varied considerably within and between the individual subtypes (M4/5 > M2/1). Using Fisher's exact test a significant difference in frequency of blast cell staining between AML and ALL was found for four MAAs (anti-lysozyme, MAC387, Ki-M1p, and KP1) and two of the three myeloid cell markers applied (Ki-My2p and anti-neutrophil elastase). Of these six antibodies, the combination of anti-lysozyme and KP1 can be recommended for use in routine diagnostics for the differentiation of AML from ALL on the basis of immunohistochemical staining because both of these antibodies were found to stain a relatively large percentage of cases of AML but none of ALL. However, none of the MAAs were found to discriminate reliably between the FAB M4/5 and M1/2 subtypes of AML.