Characterization of a panel of neurofilament antibodies recognizing N-terminal epitopes.
Peptides corresponding to sequences from the amino-terminal "head" regions of the low, middle, and high molecular weight neurofilament proteins (NF-L, NF-M, and NF-H) were synthesized by a modification of the Merrifield solid-phase method, and a panel of polyclonal antibodies to these epitopes were prepared in rabbits by the injection of synthetic peptides conjugated to the carrier protein keyhole limpet hemocyanin (KLH). An additional, monoclonal antibody recognizing both glial fibrillary acidic protein (GFAP) and vimentin was also produced, by fusion of cells of the mouse myeloma line NS-1 with spleen cells from a mouse immunized with cytoskeletal extracts. Antibody specificities were confirmed by a combination of Western blotting against cytoskeletal extracts and immunofluorescence using both rat brain sections and fibroblasts transfected with fully encoding cDNAs for each neurofilament protein, driven by viral promoters.
Kaplan
M P
MP
Department of Pathology, Columbia University College of Physicians and Surgeons, New York, NY 10032.
Chin
S S
SS
Macioce
P
P
Srinawasan
J
J
Hashim
G
G
Liem
R K
RK
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000302:isolation & purification; D000911:Antibodies, Monoclonal / Q000302:isolation & purification; D015153:Blotting, Western; D002531:Cerebellum / Q000166:cytology*; D002846:Chromatography, Affinity; D003599:Cytoskeleton / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000032:analysis*; D005347:Fibroblasts / Q000166:cytology; D005455:Fluorescent Antibody Technique; D007382:Intermediate Filaments / Q000648:ultrastructure*; D051379:Mice; D008807:Mice, Inbred BALB C / Q000276:immunology; D008969:Molecular Sequence Data; D016900:Neurofilament Proteins / Q000032:analysis* / Q000138:chemical synthesis / Q000276:immunology; D009900:Optic Nerve / Q000737:chemistry*; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology; D011817:Rabbits / Q000276:immunology; D051381:Rats; D011919:Rats, Inbred Strains; D014162:Transfection
D000906:Antibodies; D000911:Antibodies, Monoclonal; D000939:Epitopes; D016900:Neurofilament Proteins; D010455:Peptides