Application

ELISA (1:1000)Frozen Sections (1:1000, see comments)Immunoblotting (1:1000)Immunocytochemistry (1:1000, see comments)Paraffin Sections (1:1000, heat pre-treatment required, see comments)Immunoprecipitation (see comments)

General description

This PhosphoDetect™ Anti-Neurofilament H Mouse mAb (SMI-31) is validated for use in ELISA, Frozen Sections, Immunoblotting, ICC, Paraffin Sections, IP for the detection of Neurofilament H.

Mouse monoclonal antibody supplied as purified antibody. Recognizes the ~180-200 kDa phosphorylated neurofilament H protein.

Recognizes the ~180-200 kDa phosphorylated neurofilament H protein in rat central nervous system (CNS) cytoskeletal preparations. Also weakly recognizes phosphorylated NF-M protein.

Immunogen

homogenized, hypothalami from Fischer 344 rat brain

Legal Information

CALBIOCHEM is a registered trademark of Merck KGaA, Darmstadt, Germany

Other Notes

Raina, A.K., et al. 1999. Neuroreport10, 1355.Yang, C.C., et al. 1998. Brain121, 1089.Giasson, B.I and Mushynski, W.E. 1996. J. Biol. Chem.271, 30404.Mirabella, M., et al. 1996. J. Neuropath. Exp. Neurol.55, 774.Xiao, J. and Monteiro, M.J. 1994. J. Neurosci.14, 1820.

Strongly recognizes phosphorylated neurofilament H and, to a lesser extent, phosphorylated neurofilament M. By immunocytochemistry this antibody broadly stains thick and thin axons and some dendrites, such as basket cell dendrites, but not Purkinje cell dendrites. Does not generally stain nerve cell bodies or other cells and tissues except peripheral axons. May also stain neuronal cell bodies in pathological conditions. Aberrant phosphorylation of neurofilament H in cell bodies can be demonstrated in neuronal cell cultures following treatment with agents that induce stress-activated protein kinase. This antibody is reported to co-immunoprecipitate neurofilament-associated kinase (NAK 115) via interaction of the antibody with the tail domain of neurofilament H. Phosphatase treatment of tissue sections or immunoblotting samples abolishes antibody reactivity. Reactivity is unaffected by trypsin treatment of samples. Tissues and cultured cells can be fixed with a variety of paraformaldehyde- or formaldehyde-containing fixatives, including Bouin′s fixative. Post-fixation in cold methanol or methanol/hydrogen peroxide facilitates access of the antibody to the epitope in frozen sections or thick tissue sections fixed in 4% paraformaldehyde and in cultured cells. For staining formalin-fixed, paraffin sections it is recommended that the de-paraffinized tissue be autoclaved in dH₂O for 10 min or boiled in Tris-buffered saline, pH 9.0, in a microwave for 15 min to expose the epitope. Antibody should be titrated for optimal results in individual systems.

Packaging

100 µL in Glass bottle

Physical form

In PBS and Thimerosal

Warning

Toxicity: Standard Handling (A)