Abdalla S, Lother H, El Missiry A, Langer A, Sergeev P, El Faramawy Y, Quitterer U.
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2009 Mar 6;284(10):6554-65.
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REAGENTS/MATERIAL: Immunoblotting and immunohistochemistry: The following antibodies were used for immunoblotting, affinity purification and immunohistochemistry:
affinity-purified rabbit/rat polyclonal anti-AT2 antibodies (raised against an antigen encompassing amino acids 320-349 of the human AT2 receptor);
affinity-purified rabbit polyclonal anti-AT2 antibodies (raised against an antigen encompassing amino acids 16-35 of the human or mouse AT2 receptor);
affinity-purified rabbit/rat polyclonal anti-M1 antibodies (raised against an antigen encompassing amino acids 231-350 of the human M2 receptor);
affinity-purified rabbit polyclonal anti-transglutaminase antibodies (raised against an antigen encompassing amino acids 1-20 of mouse transglutaminase-2);
affinity-purified rabbit polyclonal anti-Gαq/11 antibodies (raised against the C-terminus of Gαq/11). Protein detection in immunoblot: Aβ in the prefrontal cortex specimens indicative of Aβ plaque load was assessed by immunoblot with Aβ-specific rat polyclonal antibodies. Affinity-purified antibodies or F(ab)2 fragments of the respective antibodies pre-absorbed to human and mouse proteins,
respectively, were used for detection of AT2, AT1 receptors, and Gαq/11. Immunohistochemistry, radio-immunohistochemistry and immunofluorescence:
Immunohistochemical staining of transglutaminase-2, AT2 receptor, and M1 receptor were performed with F(ab)2 fragments
of pre-absorbed affinity-purified polyclonal antibodies. Sections were stained for
Aβ plaques with monoclonal antibodies crossreacting with residues 1-12
of Aβ (BAM-10) (Sigma Aldrich). Phosphorylated tau was visualized with AT8 antibodies (Pierce). Immunohistochemistry with antibodies to MAP2 (Sigma Aldrich) was used as a marker of neuronal cell bodies and dendrites.
Lentivirus-driven expression of HA transglutaminase-2 was visualized by radio-immunohistochemistry
applying anti-HA antibodies (Santa Cruz).
Neuronal cell bodies of CA1 were quantified by [125I]-labeled NeuN antibodies.