A mild anemia (hemoglobin 9 g/dl) was found in a patient from Seville (Spain) with marked morphological abnormalities in the peripheral blood smear. The red cell osmotic fragility showed a mild resistance curve with a mean cell fragility (MCF) of 0.375% NaCl (normal = 0.450). Chemical and thermal instability test and search for inclusion bodies gave positive results. Hemoglobin electrophoresis at pH 8.9 revealed absence of Hb A, a major component of fast mobility (94%), and increased Hb F and Hb A2 levels (1.5% and 4.6%, respectively). The fast fraction, isolated and purified by means of cellulose acetate electrophoresis, precipitated in acid acetone and treated with urea 8 M and mercaptoethanol, revealed an anomalous β chain. Trypsindigested globin peptides were separated by high-voltage electrophoresis at pH 6.4 and ascendant chromatography. With differential staining, an extra peptide was detected in an unusual site, more anodic than α Tp4 but in lower position. Peptide map of the fast β chain, stained with ninhydrin, and also for Tyr, confirmed the position of the new peptide and the absence of the usual β Tp13. The new peptide, separated by high-voltage electrophoresis at pH 3.5, reveled absence of Val and the presence of an additional Glu residue, which should appear only in position β 126. The diagnosis of Hb Hofu (α2β2 126 Val→Glu; H4) was reached, thus interpreting its increase and the absence of Hb A, as an association with β°-thalassemia, producing a mild hemolytic anemia. Evidence was obtained that Hb Hofu is a mild unstable hemoglobin variant.