The Immunogen: Neurofilaments are the 10nm or intermediate filament proteins found specifically in neurons, and are composed predominantly of three major proteins called NF-L, NF-M and NF-H. NF-H is the neurofilament high or heavy molecular weight polypeptide and runs on SDS-PAGE gels at 200-220 kDa, with some variability across species boundaries. Antibodies to NF-H are useful for identifying neuronal cells and their processes in tissue sections and in tissue culture. NF-H antibodies can also be useful in the diagnostics of neurofilament accumulations seen in many neurological diseases, such as Amyotrophic Lateral Sclerosis (also known as Lou Gehrig's disease) and Alzheimer's disease. MCA-9B12 is one of numerous antibodies which reacts preferentially with the axonal phosphorylated forms of NF-H. Interestingly these phosphorylated forms of NF-H are normally restricted to axons, while less phosphorylated forms are found in dendrites. However in numerous damage and disease states, phosphorylated NF-H can be detected with MCA-9B12 in dendritic and perikaryal neurofilaments.

Left: Strip blots of crude rat spinal cord extract stained with three different antibodies to phosphorylated NF-H, MCA-NAP4 (lane 1), MCA-AH1 (Lane 2) and MCA-9B12 (lane 3). All three antibodies bind to a prominent band with an apparent SDS-PAGE molecular weight of 200kDa. Right: Mixed neuron/glial cultures stained with MCA-9B12 (red) and rabbit GFAP antibody RPCA-GFAP (green). Axonal profiles are stained in red, while astrocytic cells are revealed in green. Nuclei are revealed with a fluorescent DNA stain (blue).

Antibody characteristics: MCA-9B12 is a mouse monoclonal antibody raised against a preparation of native NF-H purified from bovine spinal cord. MCA-9B12 is an IgG2b class antibody with a k light chain. It recognizes phosphorylated NF-H KSP (lysine-serine-proline) type sequences. In some species there is some cross-reactivity with the related phosphorylated KSP sequences found in the related neurofilament subunit NF-M. The antibody recognizes NF-H strongly in all mammals tested to date and also in chicken. It recognizes neurofilaments in frozen sections in tissue culture and in formalin fixed sections.

Suggestions for use: The ascites solution has a high titre and can be used at dilutions of at least 1:1,000 in immunofluorescence experiments. In western blotting using chemiluminescence it can be used at dilutions of 1:10,000 or lower.

Limitations: This product is for research use only and is not approved for use in humans or in clinical diagnosis.

References:

1.  Harris, J., Ayyub, C. and Shaw G. A molecular dissection of the carboxyterminal tails of the major neurofilament subunits NF-M and NF-H. J Neurosci Res 30:47-62 1991.

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