Production of monoclonal antibodies against neurofilament-associated proteins: demonstration of association with neurofilaments by a coimmunoprecipitation method.
A panel of monoclonal antibodies (MAbs) was produced against mouse brain proteins that bind to the tail domain of the neurofilament (NF) heavy (200-kDa) subunit (NF-H) in vitro. An in vivo association of the MAb ligands with NFs was confirmed by examining reactivity of the MAbs with immunoprecipitated NF-H complexes. Using this method we demonstrated association of the ligands of three of the MAbs with NFs. In contrast, glial fibrillary acidic protein and an unknown 97-kDa brain protein were not associated with NFs by this criterion. An 80-kDa doublet that coimmunoprecipitated with NF-H complexes, recognized by MAb 223, was shown by immunocytochemistry and immunoblotting to be synapsin Ia and Ib. Using a complementary approach, we confirmed an association of synapsin with NFs by demonstrating that immunoprecipitated synapsin I complexes contained NF-H and NF medium (160-kDa) subunits. MAbs 63 and 105 recognized a more complex set of proteins that had predominantly synaptic localizations. These data suggest that NFs may provide important support for attachment and/or transport of synaptic proteins in brain.
Starr
R
R
Medical Biotechnology Center, University of Maryland Biotechnology Institute, Baltimore.
Xiao
J
J
Monteiro
M J
MJ
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D001921:Brain / Q000378:metabolism; D015151:Immunoblotting; D007150:Immunohistochemistry; D008024:Ligands; D051379:Mice; D009419:Nerve Tissue Proteins / Q000378:metabolism; D016900:Neurofilament Proteins / Q000276:immunology* / Q000378:metabolism; D011233:Precipitin Tests / Q000379:methods*; D016704:Synapsins / Q000276:immunology / Q000378:metabolism; D014018:Tissue Distribution
D000911:Antibodies, Monoclonal; D008024:Ligands; D009419:Nerve Tissue Proteins; D016900:Neurofilament Proteins; D016704:Synapsins; C051774:neurofilament protein H