New ELISA approach based on coiled-coil interactions

作者：Liberelle Benoit; Bartholin Laurence; Boucher Cyril; Murschel Frederic; Jolicoeur Mario; Durocher Yves; Merzouki Abderrazzak; De Crescenzo Gregory^*

来源：Journal of Immunological Methods, 2010, 362(1-2): 161-167.

DOI：10.1016/j.jim.2010.09.027

• EGF
• E/K coiled-coil interactions
• ELISA test

The de novo designed heterodimeric E/K coiled-coil system has been previously demonstrated to be an excellent capture/dimerization system applicable to various needs in both biotechnology and pharmaceutical fields. Those include controlled protein dimerization, capture, purification and Western-blot detection. We here report the development of a new generation of ELISA test based on coiled-coil interactions for the direct quantitation of coil-tagged epidermal growth factor (EGF). The new approach was evaluated for its specificity, plate storability and reusability as well as for convenience when compared to commercially available systems. Our results show a similar affinity/sensitivity to standard capturing antibody-based ELISA systems and an improved affinity/sensitivity when compared to the commercially available Ni-NTA capture system. The E/K coiled-coil ELISA system was validated with respect to recovery, intra- and inter-assay variations. The practical working range was estimated to be between 52 and 34,000 pM. Furthermore, the storability and reusability of the plates was greater than the two aforementioned systems, suggesting that the E/K coiled-coil system is a good alternative to traditional tags such as polyhistidine for the development of ELISA tests aiming at quantitating coil-tagged proteins.

• 出版日期2010-10-31