Automated Determination of Serum α1-Antitrypsin by Antitryptic Activity Measurement

Roche, Denis; Mesner, Alexandra; Nakib, Malik Al; Leonard, Frederic; Beaune, Philippe

doi:10.1373/clinchem.2008.117002

Cited by 7 publications

(1 citation statement)

References 32 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Quantitation may be accomplished by a variety of methods, although the most commonly-used technique is immunonephelometry and more recently antitryptic activity(14). Identification of the deficient alleles can be performed by two methodologies referred to as phenotyping and genotyping.…”

Section: Introductionmentioning

confidence: 99%

Simultaneous Phenotyping and Quantification of α-1-Antitrypsin by Liquid Chromatography–Tandem Mass Spectrometry

Chen¹,

Snyder²,

Zhu

et al. 2011

Clinical Chemistry

View full text Add to dashboard Cite

Background α-1-antitrypsin (A1AT) deficiency results from a genetic disorder at two common loci. Diagnosis requires quantitation of A1AT and subsequent identification of the specific variant. The current algorithm of laboratory testing for the diagnosis of A1AT deficiency uses a combination of quantitation (nephelometry), genotyping and/or phenotyping. We developed a novel multiple reaction monitoring LC-MS/MS method for simultaneous quantitation of A1AT and the identification of the two most common deficiency alleles present in 95% of the patients with A1AT deficiency. Method Serum samples (n=40) were digested with trypsin and appropriate 13C/15N-labeled standard peptides added. LC-MS/MS analysis was performed with a 0.5×150 mm C18 column and H2O:acetonitrile:n-propanol (A:98/1/1/0.2 and B:10/80/10/0.2; 12 μL/min) mobile phase in positive ion mode on a TSQ Quantum triple quadrupole MS system. The A1AT concentration was obtained by comparison to a calibration curve and the phenotype by the presence or absence of variant peptides. Results were compared to the current phenotyping assay by isoelectric focusing (IEF) and the immunonephelometry quantitative assay. Results For the A1AT allele detection, in 39 of 40 samples, the LC-MS/MS results were identical to those obtained by IEF gel electrophoresis. The single discrepant result was rerun by IEF at a lower dilution and the results were in concordance. The A1AT quantitation by LC-MS/MS also compared favorably with nephelometry. Conclusion This LC-MS/MS method correlates well with current phenotyping and nephelometric assays. It is a promising method with the potential to improve the laboratory diagnosis of genetic A1AT deficiency.

show abstract

Section: Introductionmentioning

confidence: 99%