The three known mammalian Hh proteins, Sonic (Shh), Desert (Dhh) and Indian (Ihh) are structurally related and share a high degree of amino-acid sequence identity (e.g., Shh and Ihh are 93% identical). The biologically active form of Hh molecules is obtained by autocatalytic cleavage of their precursor proteins and corresponds to approximately the N-terminal one half of the precursor molecule. Although Hh proteins have unique expression patterns and distinct biological roles within their respective regions of secretion, they use the same signaling pathway and can substitute for each other in experimental systems. Sonic Hedgehog, which is expressed only during embryogenesis, is instrumental in patterning the early embryo. It has been implicated as the key inductive signal in patterning of the ventral neural tube, the anterior-posterior limb axis, and the ventral somites. Of three human proteins showing sequence and functional similarity to the Sonic Hedgehog protein of Drosophila, this protein is the most similar.
Applications
Suitable for use in ELISA, Western Blot, Immunohistochemistry and Neutralization. Other applications not tested.
Recommended Dilution
ELISA (Capture): 2-8ug/ml Immunohistochemistry: 8-25ug/ml using perfusion fixed frozen sections of mouse brain and embryos. Western Blot: 2ug/ml on human and mouse brain and stomach tissue Neutralization: Measured by its ability to neutralize sonic hedgehog alkaline phosphatase production in C3H10TT1/2 mouse embryonic bibroblast cell line. ND50 is typically 5-15ug/ml in the presence of 5ug/ml recombinant mouse sonic hedgehog N-terminus. Optimal dilutions to be determined by the researcher.
Storage and Stability
Lyophilized and reconstituted products are stable for 12 months after receipt at -20°C. Reconstitute with sterile PBS. Aliquot to avoid repeated freezing and thawing. Store at -20°C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.
Immunogen
Recombinant protein corresponding to Cys25-Gly198 (Lys122Arg) from mouse sonic hedgehog, expresssed in E. coli.
Form
Supplied as a lyophilized powder from PBS, pH 7.4, 5% trehalose. Reconstitute with 200ul/1ml sterile PBS.
Purity
Purified by Protein G affinity chromatography from hybridoma culture supernatant. |Endotoxin: ≤0.1EU/ml (LAL)
Specificity
Recognizes recombinant mouse Shh N-terminal peptide. In Western Blot, ~50% crossreactivity with the N-terminal peptide from recombinant mouse Dhh (aa 23-198), no cross-reactivity with recombinant mouse Ihh (aa 66-240), and no crossreactivity with the C-terminal peptides of recombinant mouse Dhh (aa199-396) or recombinant mouseShh (aa199-437). Species Crossreactivity: human.