title: Direct measurement of glycine-extended adrenomedullin in plasma and tissue using an ultrasensitive immune complex transfer enzyme immunoassay in rats.
abstract: The mature form of the vasodilator peptide adrenomedullin (AM-m) is synthesized from a glycine-extended precursor (AM-Gly) by enzymatic amidation. We have developed a highly sensitive enzyme immunoassay (Immune Complex Transfer Enzyme Immunoassay; ICTEIA) that enables us to measure levels of AM-Gly in plasma and tissue directly. The detection limit of this assay is 1 amol/assay, and the intra- and inter-assay precision are 4.5-14.1% and 9.9-20.5%, respectively. Dilution curves for plasma samples showed good linearity, and the analytical recovery was 107-116.6%. Using ICTEIA, we determined that the plasma concentration of immunoreactive AM-Gly is substantially higher than that of AM-m (5.22 +/- 2.56 vs. 1.21 +/- 0.79 fmol/ml). In contrast, levels of AM-Gly were much lower than those of AM-m in the lung, heart, kidney, adrenal gland and liver. We also evaluated AM-Gly and AM-m levels in rats in a morbid state induced by intraperitoneal administration of lipopolysaccharide (LPS). In most tissues, levels of AM-m and AM-Gly were both increased by LPS; however, AM-Gly/AM-m ratios were not significantly affected, which suggests that AM-Gly is rapidly converted to AM-m in tissue.
authors.affiliation: First Department of Internal Medicine, Miyazaki Medical College, 5200 Kihara, Kiyotake, Miyazaki 889-1692, Japan.
authors.forename: Junichi
authors.initials: J
authors.lastname: Yamaga
authors.forename: Seiichi
authors.initials: S
authors.lastname: Hashida
authors.forename: Kazuo
authors.initials: K
authors.lastname: Kitamura
authors.forename: Mariko
authors.initials: M
authors.lastname: Tokashiki
authors.forename: Toshihiro
authors.initials: T
authors.lastname: Aoki
authors.forename: Haruhiko
authors.initials: H
authors.lastname: Inatsu
authors.forename: Naoto
authors.initials: N
authors.lastname: Ishikawa
authors.forename: Kenji
authors.initials: K
authors.lastname: Kangawa
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Morishita
authors.forename: Tanenao
authors.initials: T
authors.lastname: Eto
mesh_terms: D000311:Adrenal Glands / Q000737:chemistry; D053607:Adrenomedullin; D000818:Animals; D000936:Antigen-Antibody Complex; D002138:Calibration; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods* / Q000592:standards; D005998:Glycine; D007201:Indicator Dilution Techniques; D007668:Kidney / Q000737:chemistry; D008099:Liver / Q000737:chemistry; D008168:Lung / Q000737:chemistry; D008297:Male; D009206:Myocardium / Q000737:chemistry; D010455:Peptides / Q000032:analysis* / Q000097:blood* / Q000276:immunology; D051381:Rats; D017208:Rats, Wistar; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D000936:Antigen-Antibody Complex; D010455:Peptides; D053607:Adrenomedullin; D005998:Glycine