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A monoclonal antibody (AAP1) to human intestinal alkaline phosphatase (ALP) was produced by immunizing a mouse with D98/AH-2 (HeLa) cells, which produce the enzyme ectopically. The antibody, which did not inhibit enzyme activity using p-nitrophenyl phosphate as the substrate, was of the IgG2A class and did not show complement-dependent cytotoxicity. In trace binding assays AAP1 bound only to cells that expressed an intestinal-like form of human ALP, including some human intraspecific (D98/AG-2 x human lymphocyte or fibroblast) hybrids. Immunoprecipitation of immune complexes from cell-free extracts of D98/AG-2 cells, using protein A containing S. aureus and AAP1 antibody, resulted in precipitation of all the ALP activity. The precipitated material had a subunit molecular weight of 80,000 daltons, as estimated by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate. In non-denaturing conditions, AAP1 antibody prevented the migration of ALP activity into the gel when cell-free extracts were made from human adult or fetal intestine, or D98/AH-2 cells. Similarly, AAP1 could be used to precipitate ALP activity from these extracts but not from extracts of human liver, kidney or placenta.

Original publication




Journal article


Tissue Antigens

Publication Date





303 - 312


Alkaline Phosphatase, Animals, Antibodies, Monoclonal, Antibody Specificity, Binding Sites, Antibody, Cell Fusion, Cell Line, Chemical Precipitation, Female, Fetus, HeLa Cells, Humans, Intestines, Kidney, Liver, Mice, Mice, Inbred Strains, Molecular Weight, Placenta, Pregnancy