Immunohistochemical localization of human liver alcohol dehydrogenase in liver tissue, cultured fibroblasts, and HeLa cells.
Human liver alcohol dehydrogenase (ADH, EC 1.1.1.1) was purified by double ternary complex affinity chromatography on Sepharose-4-(3-[N-6 aminocaproyl]aminopropyl) pyrazole. The purified enzyme preparation still contains several isoenzymes reflecting the isoenzyme composition of the starting material. Antibodies against this mixture of isoenzymes were elicited in rabbits. The specificity of the antiserum was tested by double immunodiffusion, enzyme-linked immunosorbent assay, immunoprecipitation of ADH enzymatic activity, and adsorption to ADH, which was immobilized to Ultrogel AcA 44 by the use of glutardialdehyde as the coupling agent. Protein-A peroxidase with diaminobenzidine or amino ethyl carbazole as substrate, served to detect binding of anti-human liver ADH antibodies in human liver thin sections, cultured human skin and lung fibroblasts, and HeLa cells. Fluorescein-conjugated antibodies were also used in direct immunofluorescence on liver tissue. In the human liver, ADH was found to be localized in the cytoplasm of hepatocytes. Differences in the staining intensity of hepatocytes may reflect differences in ADH content. Strongly stained hepatocytes were localized mainly around the central veins. Perinuclear staining is often seen, especially in the more lightly stained cells. Human skin and lung fibroblasts, as well as HeLa cells, all exhibited positive staining for ADH. The pattern was identical to that found in hepatocytes, although the staining intensity was much weaker, indicating a lower ADH content.
D000429:Alcohol Oxidoreductases / Q000032:analysis*; D000906:Antibodies / Q000032:analysis; D000917:Antibody Formation; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D003593:Cytoplasm / Q000201:enzymology; D005347:Fibroblasts / Q000201:enzymology; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells / Q000201:enzymology; D006651:Histocytochemistry; D006801:Humans; D007527:Isoenzymes / Q000032:analysis*; D008099:Liver / Q000201:enzymology*; D008168:Lung / Q000201:enzymology; D012867:Skin / Q000201:enzymology
D000906:Antibodies; D007527:Isoenzymes; D000429:Alcohol Oxidoreductases