Figure 1. Schematic of experimental procedures for generation of novel anti-Sonic Hedgehog (Shh) candidate therapeutic antibodies directed at the carboxy (C)-terminal.

(A) Two synthetic peptide mimics of Shh (AA 247-264 and AA 448-462) were conjugated to carrier protein KLH and used as the immunogen. (B) Dual administration of both peptides was used to elicit an immune response in Sp2/0-Ag1 mice. (C) B cells from immunized murine lymph nodes were isolated and fused to myeloma cells for hybridoma generation. (D) Protein-based screening analyses guided the selection of the best hybridoma clones followed by one round of sub-cloning to isolate monoclonal antibody-producing hybridoma cells (E) Large-scale preparations of purified antibodies from hybdridomas were used for in vitro, in vivo, and ex vivo investigations.