PubMed | ANSES French Agency for Food, Indicia Production. and AJ Roboscreen GmbH.
Type: | Journal: Journal of visualized experiments : JoVE | Year: 2015
In addition to established methods like Western blot, new methods are needed to quickly and easily quantify disease-associated -synuclein (S(D)) in experimental models of synucleopathies. A transgenic mouse line (M83) over-expressing the human A53T S and spontaneously developing a dramatic clinical phenotype between eight and 22 months of age, characterized by symptoms including weight loss, prostration, and severe motor impairment, was used in this study. For molecular analyses of S(D) (disease-associated S) in these mice, an ELISA was designed to specifically quantify S(D) in sick mice. Analysis of the central nervous system in this mouse model showed the presence of S(D) mainly in the caudal brain regions and the spinal cord. There were no differences in S(D) distribution between different experimental conditions leading to clinical disease, i.e., in uninoculated and normally aging transgenic mice and in mice inoculated with brain extracts from sick mice. The specific detection of S(D) immunoreactivity using an antibody against Ser129 phosphorylated S by ELISA essentially correlated with that obtained by Western blot and immunohistochemistry. Unexpectedly, similar results were observed with several other antibodies against the C-terminal part of S. The propagation of S(D), suggesting the involvement of a prion-like mechanism, can thus be easily monitored and quantified in this mouse model using an ELISA approach.