Abdalla S, Lother H, El Missiry A, Langer A, Sergeev P, El Faramawy Y, Quitterer U. Angiotensin II AT2 receptor oligomers mediate G-protein dysfunction in an animal model of Alzheimer disease. J Biol Chem. 2009 Mar 6;284(10):6554-65. PubMed Abstract

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-Gα_q/11 antibodies (raised against the C-terminus of Gα_q/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 Gα_q/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.