Methods for the production of multimeric immunoglobulins, and related compositions

BRIEF DESCRIPTION OF THE DRAWINGS

0079 FIG. 1 provides a listing of exemplary proteins for use in the heteromultimeric-fusion-proteins and heteromultimeric-protein-complexes provided herein.

0080 FIG. 2 Coomassie stained protein gel showing the partitioning of assembled antibody complexes with the oil body (OB) or the soluble undematant (U) fraction from wild type (wt) Arabidopsis C24 or transgenic SBS4803 seeds. The arrow indicates the high molecular weight antibody complexes in non-reduced samples separated by SDS-PAGE as evident by the mouse IgG1 and purified D9 MAb control lanes.

0081 FIG. 3. A) Coomassie stained gel of Arabidopsis total protein extracts showing reduced or non-reduced samples from wild type (wt) seeds and transgenic SBS4809 seeds expressing chimeric heavy and light antibody chains (Lines 6 and 13). Mouse (Mm) and human (Hu) samples of IgG1 antibody are included as controls. B) Western blots showing human heavy chain IgG Fc-specifc detection and human kappa chain-specific detection. Reduced samples were separated on SDS-PAGE to identify individual antibody chains, while non-reduced samples were separated to identify antibody assemblies of heavy and light chains covalently bound by disulfide bonds. Both heavy and light chains are detected in the assembled antibody complex (non-reduced samples; arrow). The migration of this complex is comparable to the mouse and human IgG1 control protein.

0082 FIG. 4 (and SEQ ID NO:38) shows the amino acid sequence of the five immunoglobulin-binding domains in the Protein A sequence of Staphylococcus aureus.

0083 FIG. 5 (and SEQ ID NO:39) shows the DNA and encoding amino acid sequence of the Protein A insert in pSBS2904.

0084 FIG. 6. Individual wild type (wt) or transgenic safflower seeds were extracted and oil body (OB) and soluble undernatant (U) fractions were analyzed by Western blot. Detection was performed using a goat anti-human IgG Fc-specific secondary antibody (ICN Biomedicals Inc.). Seeds analyzed were from individual transgenic lines (Protein A-oleosin SBS4901, chimeric heavy and light chain SBS4810) or seeds resulting from the cross of the SBS4901 and SBS4810 transgenic lines. The double transgenic seed (SBS4810SBS4901) and single transgenic seed (SBS4810 −) resulting from the cross are compared to the single transgenic lines.