A novel and sensitive ELISA reveals that the soluble form of IL-13R-α2 is not expressed in plasma of healthy or asthmatic subjects

Background IL-13 plays a key regulatory role in asthmatic responses and immunity to parasitic infection. In vivo, IL-13R-alpha 2 is a critical modulator of IL-13 bioactivity. When inducibly expressed on the surface of fibroblasts and other cell types under inflammatory conditions, IL-13R-alpha 2 contributes to resolution of IL-13 responses. A soluble form of IL-13R-alpha 2 (sIL-13R-alpha 2) can be detected in murine circulation, and functions as a regulator of IL-13 bioactivity. In humans, sIL-13R-alpha 2 has been more difficult to detect. Recently, novel assay systems have been described to quantitate sIL-13R-alpha 2 in human circulation, and revealed unexpectedly high levels of sIL-13R-alpha 2 in healthy subjects. Objective To verify sIL-13R-alpha 2 quantitation in human plasma samples under stringent conditions of signal verification and false-positive detection. Methods A standard ELISA protocol was evaluated for specificity using false-positive detection reagents. A more stringent ELISA protocol was developed by optimizing the composition of blocking and dilution buffers. Results Using the stringent assay protocol, endogenous sIL-13R-alpha 2 was undetectable in plasma samples from a total of 120 asthmatics and 20 healthy subjects, and in bronchoalveolar lavage fluid from 10 asthmatics and eight healthy subjects undergoing allergen challenge. Conclusion These results underscore the necessity to perform rigorous assay controls in the biological matrix to be tested. Because the soluble form could not be demonstrated, our findings question a role for sIL-13R-alpha 2 in the regulation of IL-13 bioactivity, and highlight the potentially important contribution of the membrane-bound form of IL-13R-alpha 2 in humans.