Detectionof Genetic Variantsof a 1 - AntitrypsinwithSite - SpecificMonoclonal Antibodies

The serum protein a1-antitrypsin (a1-AT) serves as the major inhibitor of neutrophil elastase. The most common allele of the a.1-AT gene is designated as p1Mrn The Z mutation is a single-base substitution of the normal M allele, causing a Glu -* Lys change at position342 in the molecule. The ZZ phenotype is associated with a severe deficiency of a1-AT, serum concentrations of the protein being 10% of normal. Individualswith an a1-AT deficiency are at an increased risk of developing emphysema. To generate antibodies that specifically detect the 342 position in the context of the flanking sequences, we synthesized several peptides that included the 342 position for both the M and the Z variant. Immunization with variantspecific peptide-carrier conjugates elicited a1-AT variantspecific responses, as determined in a direct enzymelinked immunoassay. Monoclonal antibodies (MAbs) were selected with different specificity for the 342 region: MAbs F43 recognize only the a1-AT sequence with 2Glu, i.e., all variant proteins that are non-Z, either from heteroor homozygous individuals; MAbs F50 recognize only the sequence with Lys, i.e., all Z-variant proteins in ZZ or heterozygous individuals; MAbs F46 recognize a1-AT with either Lys or Glu, all variant proteins with sequences as in the peptides used. Z homoand heterozygotes were detected with our MAbs in a rapid and simple immunoblot assay. Other variants (M, S, and F) can also be assigned on the basis of the electrophoretic pattern. This sensitive detection method is very easy, rapid, and straightforward and provides a powerful tool for diagnosis of the a1-AT deficiencies, allowing early treatment (augmentation of a1-Afl and proper advice on lifestyle practices.