Production of monoclonal antibodies against inactivated alpha 1-antitrypsin. Cross-reactivity with complexed alpha 1-antitrypsin and application in an assay to determine inactivated and complexed alpha 1-antitrypsin in biological fluids.
15 different monoclonal antibodies (mcAbs) have been raised against the cleaved (inactive) form of the serpin alpha 1-antitrypsin (AT). In initial experiments these mcAbs were analysed for their ability to bind the native and the cleaved form of this inhibitor: eight of the 15 mcAbs appeared to react predominantly with cleaved AT. Additional experiments with mixtures of purified native AT, AT complexed to neutrophilic elastase and inactivated AT revealed that all mAbs that preferentially reacted with inactivated AT also bound to complexed AT. Using two of the mcAbs against inactivated AT a quantitative and sensitive sandwich-type radioimmunoassay was developed to determine levels of proteolytically inactivated AT in biological fluids. With this assay increased levels of inactivated AT were found in synovial fluid from patients with rheumatoid arthritis corresponding to about 2.4% (range 0.3-11%) of total AT. Approximately 10% of this inactivated AT appeared to consist of AT complexed to neutrophil elastase. The mcAbs described here further illustrate the structural resemblance between the complexed and cleaved forms of AT. In addition, these mcAbs appear to be useful tools for the study of AT in human disease.
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service, Amsterdam.
J J
JJ
Abbink
A M
AM
Kamp
A J
AJ
Swaak
C E
CE
Hack
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D001172:Arthritis, Rheumatoid / Q000276:immunology; D002918:Chymotrypsin / Q000494:pharmacology; D003429:Cross Reactions / Q000276:immunology; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D009504:Neutrophils / Q000201:enzymology; D010196:Pancreatic Elastase / Q000276:immunology; D011233:Precipitin Tests; D011489:Protein Denaturation / Q000187:drug effects; D011817:Rabbits; D011863:Radioimmunoassay; D013582:Synovial Fluid / Q000276:immunology; D000515:alpha 1-Antitrypsin / Q000032:analysis / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000515:alpha 1-Antitrypsin; D002918:Chymotrypsin; D010196:Pancreatic Elastase