Although protein 4.1 was originally identified as an element of the erythrocyte membrane skeleton, its presence in most mammalian cell types is now well described. Antibodies raised against erythrocyte protein 4.1 or synthetic peptides corresponding to the spectrin-actin-binding domain of protein 4.1 react with plasma membranes and, unexpectedly, nuclei of different cell types. Nuclear staining was further confirmed in isolated nuclei prepared from rat liver and human leukaemic cell lines. Immunoblot analysis of subcellular fractions derived from these cells revealed three prominent proteins, of 80, 135 and 145 kDa. The structural relationship of the high-molecular-mass proteins with erythrocyte protein 4.1 was demonstrated by peptide mapping. These results indicate that mammalian nucleated cells contain several isoforms of erythrocyte protein 4.1 and that some high-molecular-mass forms may primarily reside in the nucleus.