Inhibition of Aspergillus fumigatus elastase with monoclonal antibodies produced by using denatured elastase as an immunogen.
In preparing monoclonal antibodies to the elastase from Aspergillus fumigatus, we found that the enzyme was weakly immunogenic in BALB/c mice. Antiserum titers were only 1:1,000 to 1:5,000, and hybridomas secreted nonspecific immunoglobulin M (IgM). Denaturing the elastase in 0.5% sodium dodecyl sulfate at 80 degrees C for 10 min prior to injection increased titers of antiserum against the nondenatured (native) enzyme 10-fold. Of eight hybridomas selected following immunization with the denatured enzyme, seven produced IgG reactive with the native enzyme and one produced nonspecific IgM. The nondenatured immunogen tested again yielded mainly IgM producers. Immunoblots and enzyme-linked immunosorbent assay showed that the IgG monoclonal antibodies were reactive with both the denatured and nondenatured fungal elastases; none cross-reacted with human neutrophil elastase, porcine pancreatic elastase, or Pseudomonas elastase. Elastase-specific polyclonal antibody produced in mice inhibited elastase activity beginning at a molar ratio (antibody to elastase) of 4:1, and activity was completely inhibited at 14.5:1. Some individual monoclonal antibodies partially inhibited elastase, but certain pairs, at a molar ratio of each antibody to elastase of 5.4:1, acted synergistically to inhibit the activity completely.
Frosco
M
M
Department of Biochemistry and Microbiology, Cook College, Rutgers University, New Brunswick, New Jersey 08903.
Fahed
C
C
Chase
T
T
Macmillan
J D
JD
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D001232:Aspergillus fumigatus / Q000201:enzymology*; D005260:Female; D007114:Immunization; D051379:Mice; D008807:Mice, Inbred BALB C; D010196:Pancreatic Elastase / Q000037:antagonists & inhibitors / Q000276:immunology*; D011489:Protein Denaturation
D000911:Antibodies, Monoclonal; D010196:Pancreatic Elastase