Scientific Resources

Flow cytometry-based immunoassays for determining functional levels of plasma C1 inhibitor for the diagnosis of hereditary angioedema (HAE)


To be presented by Kusumam Joseph, PhD at the 2023 American Academy of Allergy, Asthma, and Immunology Meeting in San Antonio, TX, 2/26/2023.


HAE types I and II are caused by a deficiency of functional C1 inhibitor (fC1-INH), leading to overproduction of bradykinin. Specific and sensitive methods to quantitate fC1-INH are required for accurate and timely diagnosis of HAE. Two commercially available assays either measure residual C1-esterase (C1s) activity or a complex ELISA measuring activated C1s. More physiologically relevant alternative methods measuring functional binding of Factor XIIa or kallikrein have been reported but they are not commercially available.


Assays were developed using biotinylated factor XIIa, kallikrein, or C1s bound to streptavidin-coated microbeads. Incubation with plasma was followed by the detection of bound C1-INH. The assays were compared with ELISA-based assays and with results from a commercial lab.


After standard curves were developed for quantification of
C1-INH, serial dilutions of normal plasma were employed to validate the ability to detect a known concentration of C1-INH in the plasma as a percent of normal. Thirty-seven samples each from normal and HAE types I and II subjects were then tested. The level of fC1-INH in all HAE plasma tested was less than 40% of our normal control. This was evident regardless of whether we measured factor XIIa–C1-INH, kallikrein–C1-INH, or C1sC1-INH complexes, and all three assays were in agreement. By contrast,
results obtained from a commercial lab revealed equivocal (41-67%) or normal (>67%) levels of fC1-INH in 6/37 samples tested.


The results indicate that the new flow cytometry-based
assays are specific, sensitive, and easy to perform for the diagnosis of HAE.