Characterization of HPC-1 antigen, an isoform of syntaxin-1, with the isoform-specific monoclonal antibody, 14D8.
We raised polyclonal and monoclonal antibodies against rat recombinant HPC-1/syntaxin 1A lacking a transmembrane domain. The polyclonal antibody recognized two major bands at 35 and 40 kDa from rat brain membranes. A hybridoma clone designated 14D8, however, recognized only one band at 35 kDa. A polyclonal antibody detected recombinant syntaxin 1B, as well as HPC-1/syntaxin 1A on an immunoblot, whereas 14D8 recognized recombinant HPC-1/ syntaxin 1A, but not syntaxin 1B. Therefore, 14D8 is specific for HPC-1/syntaxin 1A. Using this monoclonal antibody, we investigated the expression of HPC-1/syntaxin 1A in the rat hippocampal membranes. HPC-1/syntaxin 1A was present even in the embryonic d 19 ( E19) hippocampal membranes, and it increased during the next two postnatal wk. Pyramidal cell axons were intensely stained with the 14D8 monoclonal antibody, suggesting that HPC-1/syntaxin 1A was not restricted to the presynaptic terminal. Furthermore, we investigated the phosphorylation of HPC-1/syntaxin 1A in the rat brain membranes. HPC-1/syntaxin 1A affinity-purified on a 14D8 IgG-coupled column was recognized by antiphosphoserine antibody, but not by antiphosphotyrosine and phosphothreonine antibodies.[1]References
- Characterization of HPC-1 antigen, an isoform of syntaxin-1, with the isoform-specific monoclonal antibody, 14D8. Kushima, Y., Fujiwara, T., Sanada, M., Akagawa, K. J. Mol. Neurosci. (1997) [Pubmed]
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