The carboxy-terminal amino acid sequence of the soluble form of the 53,000 mol. wt monocyte surface antigen, CD14, was determined by carboxypeptidase Y digestion and compared with the complete amino acid sequence of this protein as predicted from the structure of cloned cDNA [Goyert et al. Science 239, 497-500 (1988)]. The soluble antigen isolated from urine appears to lack eight C-terminal amino acid residues predicted for the full-size translation product, but possesses a major part of the C-terminal hydrophobic domain originally suggested as the membrane-spanning segment. The CD14 antigen can be removed from the monocyte surface by phosphatidylinositol-specific phospholipase C treatment, indicating that this glycoprotein is anchored in the membrane by a phospholipid and is not a transmembrane protein. The soluble form occurring in serum and in supernatants of cultured monocytes thus probably arises by phospholipase-mediated cleaving off the cell surface antigen. A sensitive sandwich ELISA was developed using a monoclonal anti-CD14 antibody, MEM-18, and polyclonal rabbit anti-CD14 antiserum for quantitation of the soluble antigen concns in sera and cell culture supernatants. Using this assay, the antigen present in the supernatant of phospholipase treated peripheral blood mononuclear cells could be estimated. The assay was also used for estimation of the concns of the soluble form of the CD14 antigen in human sera.
- MeSH
- antigeny povrchové * analýza MeSH
- elektroforéza v polyakrylamidovém gelu MeSH
- fosfolipasy typu C farmakologie MeSH
- lidé MeSH
- molekulární sekvence - údaje MeSH
- monocyty imunologie MeSH
- rozpustnost MeSH
- sekvence aminokyselin MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- antigeny povrchové * MeSH
- fosfolipasy typu C MeSH
A monoclonal antibody MEM-43 was prepared, which recognizes an antigen expressed on all peripheral blood leucocytes, on erythrocytes and several cell lines, but is absent from U937, Nalm-6, Daudi and Raji cell lines. The antigen isolated by immunoaffinity chromatography from several cell lines is an 18,000-25,000 mol. wt glycoprotein. An apparently identical antigen isolated from erythrocytes binds to several lectins and has a 14,000 mol. wt polypeptide backbone, modified by an endoglycosidase F-sensitive carbohydrate moiety. The epitope recognized is reduction-sensitive. The sequence of N-terminal 17 amino acid residues was determined; five out of six N-terminal amino acids are identical to those found at the N-terminus of the mouse lymphocyte surface antigen Ly-6C. The antigen is completely released from the cell surface after treatment with phosphatidylinositol-specific phospholipase C.
- MeSH
- antigeny povrchové imunologie metabolismus MeSH
- buněčná membrána imunologie metabolismus MeSH
- buněčné linie MeSH
- elektroforéza v polyakrylamidovém gelu MeSH
- fosfatidylinositoly metabolismus MeSH
- glykoproteiny imunologie metabolismus MeSH
- leukocyty imunologie MeSH
- lidé MeSH
- molekulární sekvence - údaje MeSH
- monoklonální protilátky imunologie metabolismus MeSH
- myši MeSH
- sekvence aminokyselin MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- antigeny povrchové MeSH
- fosfatidylinositoly MeSH
- glykoproteiny MeSH
- monoklonální protilátky MeSH
