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There are potentially two binding sites for C1q on IgG, one on each CH2 domain of the gamma heavy chains, close to the lower hinge region. It is not clear whether the presence and involvement of both the C1q binding sites is necessary to induce the activation signal of human IgG. In order to clarify this issue, we made a hybrid mutant IgG1/IgG3 molecule where the IgG1 half of the molecule was made unable to activate complement through the introduction of a P329A mutation. The IgG3 half of the molecule was mutated to harbor a hinge region identical to that of IgG1, and for detection a peptide tag derived from p21ras was introduced into the FG loop of the CH1 domain. The hybrid IgG1P329A/IgG3h1‐ras molecules were isolated by Protein A affinity chromatography and shown to activate complement and induce complement‐mediated lysis at the same levels as wild‐type IgG1 and IgG3h1‐ras molecules. Thus, one C1q binding site per IgG is sufficient to induce activation. Wild‐type human IgG molecules might also normally expose only one C1q binding site as already shown for interaction with FcγR, were IgG expose one binding site per molecule.

In order to prevent or ameliorate autoimmune disease, it would be desirable to induce central tolerance to peripheral self-antigens. We have investigated whether recombinant antibodies (Ab) that deliver T cell epitopes to antigen-presenting cells (APC) in the thymus can be used to induce thymocyte deletion. Troybodies are recombinant Ab with V regions specific for APC surface molecules that have T cell epitopes genetically introduced in their C domains. When MHC class II-specific Troybodies with the k2 315 T cell epitope were injected into k2 315 -specific TCR transgenic mice, a profound deletion of CD4 + 8 + thymocytes was observed. MHC class II-specific Troybodies were 10-100-fold more efficient than non-targeting peptide Ab, and 500-fold more efficient than synthetic peptide at inducing deletion. Similar findings were observed when MHC class II-specific Troybodies with the OVA 323-339 T cell epitope were injected into OVA-specific TCR transgenic mice. Although deletion was transient after a single injection, newborn mice repeatedly injected with MHC class II-specific Troybodies for 4 weeks, had reduced antigen-specific T cells in peripheral lymphoid tissues and reduced T cell responses. These experiments suggest that Troybodies constructed to target specifically thymic APC could be useful tools for induction and maintenance of central T cell tolerance in autoimmune diseases.

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