A Recombinant Ig Fragment (IgCw-gek) Comprising the Cg1-Ce2-4 and Ck Domains Is an Alternative Reagent to Human IgE

Abstract

Human IgE is useful for immunological assays, such as sensitization of FcepsilonRI-positive cells and IgE measurement. In this study, we report the development of a recombinant Ig fragment, designated IgCw-gammaepsilonkappa, as an alternative reagent to human IgE. IgCw-gammaepsilonkappa ( approximately 130 kDa) comprises two hybrid constant H chain regions (Cgamma(1)-Cepsilon(2-4), each approximately 53 kDa) and two constant kappa L chains (C(kappa), each approximately 12 kDa) and lacks a V domain. The presence of Cgamma(1) instead of Cepsilon(1) within the H chain increased the production yield and facilitated assembly of the H and L chains. IgCw-gammaepsilonkappa was produced in cultured human embryonic kidney 293F cells, with a yield of approximately 27 mg/l. IgCw-gammaepsilonkappa bound to human FcepsilonRIalphaRs expressed on the surface of rat basophilic leukemia-2H3 cells. A beta-hexosaminidase release assay revealed that the biological activity of IgCw-gammaepsilonkappa was comparable with that of IgE. The IgE concentration measured using IgCw-gammaepsilonkappa as a standard was similar to that measured using IgE as a standard. These results suggest that the IgCw-gammaepsilonkappa molecule retains the basic characteristics of IgE, but does not cross-react with Ags, making it an alternative to the IgE isotype references used in a variety of immunological assays.