Production of monoclonal antibodies to prostromelysin (ProMMP-3) and establishment of a quantitative prostromelysin ELISA assay.
Human prostromelysin (59 kDa) was purified from the conditioned medium of IL-1-stimulated human dermal fibroblasts and anti-prostromelysin monoclonal antibodies were produced and identified by ELISA assay. Using prostromelysin, a C-terminally truncated recombinant form of prostromelysin consisting of amino acids 1-255, and their respective activated enzymes, we have begun mapping the epitopes recognized by these monoclonal antibodies. Various patterns of reactivity against the proenzymes and activated enzymes were observed. In further attempts to map the epitopes, we employed synthetic peptides representing hydrophilic regions of the primary amino acid sequence of prostromelysin. Our monoclonal antibodies did not recognize these peptides, suggesting that the antibodies may be recognizing conformational epitopes composed of non-linear portions of prostromelysin. Using these monoclonal antibodies, we have developed a quantitative prostromelysin sandwich ELISA assay.
Molecular Sciences Research, Hoffmann-La Roche Inc., Nutley, NJ 07110.
D H
DH
Presky
V L
VL
Wilkinson
M D
MD
Flannery
E
E
Korkmaz
R
R
Walsky
L J
LJ
Mondini-Minetti
N
N
Fotouhi
W
W
Levin
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D015153:Blotting, Western; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D004792:Enzyme Precursors / Q000276:immunology* / Q000302:isolation & purification; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D005347:Fibroblasts / Q000201:enzymology; D006801:Humans; D008666:Metalloendopeptidases / Q000276:immunology* / Q000302:isolation & purification; D051379:Mice; D008807:Mice, Inbred BALB C; D012680:Sensitivity and Specificity
D000911:Antibodies, Monoclonal; D004792:Enzyme Precursors; D008666:Metalloendopeptidases; C048047:prostromelysin