Alpha1Center uses the AlphaTest® Kit to complete as many as four levels of testing for Alpha-1:

Immunoassay (initial test performed on all samples received by Alpha1Center)

  • What is this test? An automated immunoassay is first performed to determine the concentration (or level) of alpha1-antitrypsin in the blood sample provided. The assay is a double-antibody fluorescence immunoassay that is performed in duplicate. A standard curve is included on each assay plate as well as two levels of control.
  • When is this test performed? This is the initial test performed on all samples submitted to the laboratory.
  • Why is this test performed? Alpha-1 is defined by a low concentration of alpha1-antitrypsin antitrypsin in the blood. Thus, the natural first assay is an immunoassay.

Phenotyping (second level of testing)

  • What is this test? The phenotyping procedure is isoelectric focusing of the sample in the pH range of 4.2 to 4.9. This procedure takes advantage of the fact that most of the genetically abnormal alpha1-antitrypsin variants have a slightly different electrical charge than the common normal alpha1-antitrypsin.
  • When is this test performed? This test is performed on all samples submitted to the laboratory.
  • Why is this test performed? The result from this testing most often allows the type of alpha1-antitrypsin proteins in the patient's sample to be identified without drawing additional blood. This level of testing can identify heterozygotes of alpha1- antitrypsin and can detect the presence of more rare mutations, which will need to be further evaluated at the genotype level.

Genotyping (third level of testing)

  • What is this test? The genotyping procedure is a PCR protocol, using DNA extracted from the nucleated cells in the blood sample. DNA probes are used for the identification of the most common deficiency alleles, in addition to S and Z.
  • When is this test performed? If the patient's alpha1-antitrypsin level is <11 µM (27mg/dL) and/or phenotypes as PiZ, PiS,or PiSZ, laboratory protocol dictates that a genotype is automatically performed. A genotype is also performed on samples from patients who have an unusual alpha1-antitrypsin type(s) on the phenotyping test.
  • Why is this test performed? In many cases, genotyping can unequivocally identify rare mutations that are suggested by phenotyping. Alpha1Center has a commitment to expanding their offering of testing for these rare mutations as we develop new probes and laboratory processes.

DNA Sequencing (fourth level of testing)

  • What is this test? DNA sequencing of the alpha1-antitrypsin gene is the final test that is performed only on highly-selected patient samples. The procedure involves extraction of DNA from the nucleated cells in the sample, specific amplification of the coding regions of the α1-antitrypsin gene, and then “reading” the sequence of bases in all of the amplified DNA.
  • When is this test performed? If the patient’s test result is indeterminate after the first three levels of testing, this test is performed. Reasons that it might be performed include an unexplained low concentration of α1-antitrypsin in the blood (first test above) and/or abnormal α1-antitrypsin phenotype (second test, above) with no evidence of a common mutation by genotyping (third test above). Occasionally DNA sequencing is performed if a patient has a known family history of a rare α1-antitrypsin mutation.
  • Why is this test performed? For selected unusual samples with rare α1-antitrypsin mutations, DNA sequencing is the only way to provide unequivocal test results. The results might include confirming (or not confirming) the suspected presence of Null mutations that result in the production of no α1-antitrypsin protein, and conclusively demonstrating the specific nature of a rare mutation. These unequivocal results may have important implications for both the patients and their family members.