A one-step sandwich enzyme immunoassay for human matrix metalloproteinase 8 (neutrophil collagenase) using monoclonal antibodies.
A one-step sandwich enzyme immunoassay (EIA) system for human matrix metalloproteinase 8 (MMP-8, neutrophil collagenase, EC 3.4.24.7) has been established with a pair of monoclonal antibodies prepared against the zymogen of MMP-8 purified from human neutrophils. MMP-8 in samples simultaneously reacted with both solid-phase and peroxidase-labeled antibodies. Sensitivity of this EIA system was 0.34 micrograms/l (5.7 pg/assay) and linearity was obtained between 0.5 and 500 micrograms/l (8.3-8300 pg/assay). The EIA system recognized both precursor and active forms of MMP-8 but not MMP-8 complexed with tissue inhibitors of metalloproteinases. There was no difference in the MMP-8 levels between the plasma samples from patients with rheumatoid arthritis or osteoarthritis and those from healthy subjects (median 6.2 micrograms/l, range 1.5-28 micrograms/l). However, the level in synovial fluids from patients with rheumatoid arthritis (median 345 micrograms/l, range 84-2860 micrograms/l) was shown to be higher than that from osteoarthritic patients. MMP-8 levels in human whole saliva from patients with periodontal diseases (median 282 micrograms/l, range 0-1420 micrograms/l) were also significantly higher than those from clinically healthy subjects (median 25 micrograms/l, range 0-100 micrograms/l). Immunoreactivity analyses showed that MMP-8 species in normal human plasma exists as a precursor but not as a complex form with tissue inhibitor of metalloproteinases (TIMP)-1 or TIMP-2.
Matsuki
H
H
Biopharmaceutical Department, Fuji Chemical Industries, Ltd., Toyama, Japan.
Fujimoto
N
N
Iwata
K
K
Knäuper
V
V
Okada
Y
Y
Hayakawa
T
T
D000818:Animals; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D001172:Arthritis, Rheumatoid / Q000097:blood / Q000201:enzymology; D015415:Biomarkers; D002460:Cell Line; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D017364:Collagenases / Q000032:analysis* / Q000097:blood / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006801:Humans; D007124:Immunoenzyme Techniques; D007150:Immunohistochemistry; D020784:Matrix Metalloproteinase 8; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight; D009504:Neutrophils / Q000201:enzymology; D010003:Osteoarthritis / Q000097:blood / Q000201:enzymology; D010920:Placenta / Q000201:enzymology; D011247:Pregnancy; D012463:Saliva / Q000201:enzymology; D018709:Statistics, Nonparametric; D013274:Stomach Neoplasms / Q000201:enzymology / Q000473:pathology / Q000601:surgery; D013582:Synovial Fluid / Q000201:enzymology; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D015415:Biomarkers; D017364:Collagenases; D020784:Matrix Metalloproteinase 8