POLYMERASE CHAIN REACTION-MEDIATED SITE-DIRECTED MUTAGENESIS DETECTION OF Z-ALPHA-1-ANTITRYPSIN AND S-ALPHA-1-ANTITRYPSIN ALLELES IN FAMILYMEMBERS

Alpha-1-antitrypsin (A1AT) deficiency is an autosomal hereditary disor der with a reduction in serum A1AT levels. In a large family, we used a polymerase chain reaction (PCR)mediated, site-directed mutagenesis a ssay to detect the two most common A1AT deficient variants, Z and S. B y coamplification, using primers for both the Z and S mutations, we we re able to detect heterozygous and homozygous genotypes for both mutat ions in a single reaction. We compared our results with phenotype stud ies obtained by standard immunofixation and isoelectric focusing techn iques at two reference laboratories. Whereas PCR and isoelectric focus ing agreed completely, there were five discrepancies in the results ob tained by the immunofixation procedure. The reference laboratory that provided these discrepant results later informed us of a quality contr ol problem that accounted for their error. The family study included 1 2 individuals representing three generations. Two individuals were MM homozygotes, three were MZ heterozygotes, four were MS heterozygotes, and three were SZ heterozygotes. A thirteenth family member was diagno sed as a ZZ homozygote at another institution. We have shown that this PCR coamplification technique provides accurate information about the M, S, and Z alleles that is at least as useful as current reference l aboratory methodologies. (C) 1996 Wiley-Liss, Inc.