Measuring Mutant and Wild-Type A1AT Proteins in Heterozygous Patients


Alpha-1 antitrypsin (A1AT) is a serine protease inhibitor that protects the lung from enzymatic damage. A1AT deficiency is an inherited disorder, which results from mutations in the A1AT gene. While more than 100 alleles have been identified, the M allele encodes the wild-type protein, and the Z and S alleles are the most common deficiency alleles.

Diagnosis of A1AT deficiency requires identification of the deficiency alleles and quantitation of circulating A1AT. Traditionally, laboratory analysis of A1AT deficiency involves immunoquantitation of total A1AT and detection of the disease-associated A1AT alleles. Standard laboratory methods for identification of A1AT alleles include isoelectric focusing of serum proteins or molecular analysis of white blood cell DNA. However, these methods don’t allow for quantitation of allelic expression.

In a recent study published in the Respiratory Research journal, Mayo Clinic researchers utilized liquid chromatography and tandem mass spectrometry (LC-MS/MS) analysis of A1AT tryptic peptides to differentiate between mutated (S and Z) and wild-type (non-S and non-Z) proteins, which allowed for quantitation of circulating allelic expression in heterozygous patients.

Leslie Donato, Ph.D.
Leslie Donato, Ph.D.

“Current methods for allele identification are not quantitative. As a result, none of the available laboratory methods were designed to quantitate the expression of the different alleles,” said Leslie Donato, Ph.D., Consultant in the Cardiovascular Laboratory Medicine and Clinical Core Laboratory at Mayo Clinic and first author on this study. “With the advent of using mass spectrometry for allele detection, we realized the method could be further developed to also measure the concentration of the alleles in heterozygous patients.”

Most patients that display symptoms of A1AT deficiency express very low circulating A1AT protein as a result of expression from two deficiency alleles. However, several studies have suggested that some patients who are heterozygous for an A1AT-deficiency allele may be at risk for clinical manifestations of the disease.

“To be honest, we don’t know whether quantitation of the individual alleles is important. We hypothesized that the variability of risk for clinical disease in heterozygous patients may be related to how much wild-type and/or mutant protein they produce,” added Dr. Donato.

The study utilized the novel assay developed at Mayo Medical Laboratories for mutant allele detection based on mass spectrometry technology. Using LC-MS/MS, the laboratory can identify the presence of the two most common deficiency alleles (the Z and S alleles) with great accuracy.

To carry out the study, the authors modified the currently available LC-MS/MS assay to be able to quantitate the concentration of the individual alleles. The results of the study indicate that heterozygous patients express a wide range of mutant to wild-type protein ratios.

“Using LC-MS/MS, we were able to determine that some heterozygous patients express the wild-type protein at concentrations that could theoretically place them at higher risk for clinical manifestations of A1AT deficiency,” said Dr. Donato.

This study shows, for the first time, that it is possible to measure mutant and wild-type A1AT proteins in heterozygous patients. Future studies will determine whether allele-specific quantitation can be used to identify heterozygous patients at greatest risk who would benefit from more intensive medical management.

“While we haven’t yet answered the question of whether allele-specific quantitation is clinically relevant, we now have the tools to address this hypothesis and continue pursuing it in future clinical studies,” said Dr. Donato.

The clinical utility of A1AT alleles quantitation is currently under investigation, therefore, it is not currently orderable. However, for more information on the Alpha-1-Antitrypsin Proteotype S/Z by LC-MS/MS, Serum test, visit

Kelley Luedke

Kelley Luedke

Kelley Luedke is a Marketing Channel Manager at Mayo Clinic Laboratories. She is the principle editor and writer of Insights and leads social media and direct marketing strategy. Kelley has worked at Mayo Clinic since 2013. Outside of work, you can find Kelley running, traveling, playing with her kitty, and exploring new foods.