Glutaraldehyde-Modified Recombinant Fel d 1: A Hypoallergen With Negligible Biological Activity but Retained Immunogenicity

Fel d 1 was cloned as a single construct linking both chains of the molecule and expressed in Escherichia coli and Pichia pastoris. After physicochemical purification, recombinant (r) Fel d 1 was chemically modified using glutaraldehyde. The effect of modification on immune reactivity was evaluated using radioallergosorbent test, CAP inhibition, competitive radioimmunoassay, enzyme-linked immunosorbent assay, basophil histamine release, and T-cell proliferation assays. Both natural and recombinant unmodified Fel d 1 were used as controls.rFel d 1 demonstrated similar IgE binding and biologic activity as its natural counterpart. Upon modification, IgE-binding potency decreased >1000-fold, translating into a >106-fold reduction in biologic activity assessed by basophil histamine release. In contrast, the modified recombinant did not show a decreased but even a moderately increased capacity (1.5-fold) to stimulate proliferation of T cells (P < 0.01). Finally, it induced specific IgG antibodies in rabbits that recognized the unmodified allergen.Chemical modification is a practical and highly effective approach for achieving hypoallergenicity of recombinant allergens with retained immunogenicity.