title: Purification of placental alkaline phosphatase and its monoclonal antibody.
abstract: Monoclonal antibodies (MAbs) to placental alkaline phosphatase (PAlP) were raised by using the hybridoma technique. Spleen cells from immunized mice were fused with the mouse myeloma line NS-1 using 50% polyethylene glycol and cultured in a selection medium. Antibodies were screened by the enzyme immunoassay using immobilized solid-phase antigen and anti-mouse immunogloblin Fab' (rabbit)-beta-D-galactosidase complex. Four double-cloned hybridomas were obtained. The cross-reactivity with different kinds of alkaline phosphatases of the raised MAbs was examined. At first, MAb 11-D-10, with which placenta was stained but liver and small intestine were not stained in indirect immunofluorescence, was selected. Then, the cross-reactivity of MAb 11-D-10 was further investigated by immunoblotting (Western blotting). MAb 11-D-10 reacted with PAlP but did not react with hepatic and intestinal alkaline phosphatase at all. The binding activity of 125I-labeled MAb 11-D-10 with different choriocarcinoma cell lines (SCH, BeWo, NaUCC-1, NaUCC-2, and NaUCC-3) was highest in SCH, then in the order of BeWo, NaUCC-3, NaUCC-1 and NaUCC-2, and correlated to the PAlP content of the cells (SCH greater than BeWo greater than NaUCC-3 greater than NaUCC-1 greater than NaUCC-2). Also the intensity of indirect immunofluorescence of the above cell lines with MAb 11-D-10 correlated with the PAlP content of each cell.
mesh_terms: D000469:Alkaline Phosphatase / Q000302:isolation & purification*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D002460:Cell Line; D002822:Choriocarcinoma / Q000276:immunology; D005260:Female; D005455:Fluorescent Antibody Technique; D058851:GPI-Linked Proteins; D007075:Immunoglobulin M / Q000276:immunology; D007527:Isoenzymes / Q000276:immunology / Q000302:isolation & purification*; D051379:Mice; D010920:Placenta / Q000201:enzymology*; D011247:Pregnancy
chemical_list: D000911:Antibodies, Monoclonal; D058851:GPI-Linked Proteins; D007075:Immunoglobulin M; D007527:Isoenzymes; D000469:Alkaline Phosphatase; C019839:alkaline phosphatase, placental