Congenital erythropoietic porphyria (CEP), a rare autosomal recessive inborn error of heme biosynthesis, results from reduced activity of uroporphyrinogen III synthase (URO-III-S) leading to an excessive production and accumulation of porphyrins. Various clinical and biochemical observations point to a relationship between degree of disease expression and metabolic disturbance. We investigated 20 patients with early onset of clinical symptoms of CEP and, additionally, the relatives of six patients. CEP was confirmed by porphyrinemia and porphyrinuria with dominance of uroporphyrin and its isomer I. The investigation of the immunological nature of the defective URO-III-S gene from unrelated patients with unknown mutations was possible thanks to an antibody against the human enzyme. URO-III-S concentration in erythrocytes was determined by ELISA. No signal was achieved when assaying nonimmune serum by ELISA, whereas there was a positive reaction with the serum after immunisation. Furthermore, specificity of immune sera is demonstrated by immunoprecipitation of URO-III-S activity which caused a 33% reduction of enzyme activity. Normal levels of immunoreactive enzyme protein 100 Ϯ10% of control (x ϮSD, n ϭ 12) with a reduced specific activity 15Ϯ8.5 % (x ϮSD, n ϭ 12) were found in erythrocytes from all patients, with the exception of a girl with a remarkably high enzyme concentration of 149% of controls and a very low specific activity of 4 %. In consequence, all patients had cross-reacting immunological material (CRIM)-positive mutations. CRIM-ratios varied between 3.2 and 24.5. The CRIM-positive nature of the gene defect indicated that the mutations altered the activity of URO-III-S. The different CRIM ratios implied the presence of various mutations, which is further evidence for the heterogeneity in the genetic defect found in CEP. URO-III-S activity was determined in erythrocyte lysates by a coupled enzyme assay. Erythrocyte URO-III-S activities of patients were reduced to 4Ϫ33% of the normal value (1.72 Ϯ 0.14 pkat/mg protein). An increase of urinary coproporphyrin isomer I (40Ϫ 61%, norm ϭ 17Ϫ31%) and a halved URO-III-S activity can serve as a biochemical test for asymptomatic heterozygous gene carriers of CEP.Keywords : congenital erythropoietic porphyria ; uroporphyrinogen III synthase.Congenital erythropoietic porphyria (CEP), classically known as Günther's disease, is an autosomal recessive inborn error of heme biosynthesis which results from a 80Ϫ90% decreased activity of uroporphyrinogen III synthase (URO-III-S), the fourth enzyme of the heme biosynthetic pathway. The enzyme deficiency leads to an excessive production, accumulation and excretion of predominantly type I porphyrins, which clinically induce severe cutaneous photodermatosis mostly in association with a hemolytic process. Usually, the disease becomes obvious in early childhood [1,2]. However, a few cases have been reported in patients with late onset of photosensitivity [3]. Furthermore, dual deficiency of URO-III-S and coproporphy- rinogen ...