Quantitative analysis for soluble elastin in circulation and cell culture fluids using monoclonal antibody-based sandwich immunoassay.
We have newly established 3 distinct murine monoclonal antibodies (MoAbs) against human soluble elastin by using chemically denatured immunogen isolated from human aorta; they are designated as HASG-2, HASG-30, and HASG-61-1. All of these MoAbs were highly reactive with soluble forms of native elastin in normal human serum. HASG-2 and HASG-61-1 MoAbs can recognize soluble bovine elastin as well as human antigen, but HASG-30 cannot. The sandwich enzyme-linked immunosorbent assay (ELISA) for human soluble elastin was developed with HASG-61-1 labeled with peroxidase and HASG-30 immobilized on the microplates. The circulating levels of soluble elastin in human healthy subjects (mean +/- SD; 42.9 +/- 19.9ng/mL; n = 85) could be measured with full accuracy and reproducibility, and gradually increased with aging. The positive correlation between the levels and ages was statistically significant (r = 0.581, p < 0.0001). In addition, we could also determine the concentration of tropoelastin secreted from cultured human dermal fibroblasts accurately by this ELISA. This simple assay can be utilized for the routine clinical laboratory screening of patients with arteriosclerotic vascular diseases or to accurately determine the concentrations of tropoelastin secreted from cultured human cells.
Shinohara
Tadashi
T
Internal Medicine I, National Defense Medical College, Saitama, Japan.
Okada
Makoto
M
Suzuki
Kimihiro
K
Ohsuzu
Fumitaka
F
Katayama
Masahiko
M
D000328:Adult; D000367:Age Factors; D000368:Aged; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000276:immunology*; D002417:Cattle; D049109:Cell Proliferation / Q000187:drug effects; D017077:Culture Media, Conditioned / Q000737:chemistry / Q000378:metabolism; D020405:Dermis / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D004549:Elastin / Q000032:analysis* / Q000097:blood / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005260:Female; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D006041:Goats; D006133:Growth Substances / Q000494:pharmacology; D006801:Humans; D008253:Macaca mulatta; D008297:Male; D051379:Mice; D008875:Middle Aged; D011817:Rabbits; D015203:Reproducibility of Results; D013552:Swine; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D014333:Tropoelastin / Q000032:analysis / Q000378:metabolism
D000911:Antibodies, Monoclonal; D017077:Culture Media, Conditioned; D006133:Growth Substances; D016212:Transforming Growth Factor beta; D014333:Tropoelastin; D004549:Elastin