Immunohistochemistry was performed on 5-μm thick sections from formalin-fixed paraffin embedded tissue. Glass-mounted sections were de-parafinized in xylene and rehydrated in ethanol and dH2O. Rabbit polyclonal antibodies used for DPR immunohistochemistry were as follows: poly-GA (Rb9880), poly-GP (Rb5823), poly-GR (Rb7810) (Leonard Petrucelli, Mayo Clinic, Jacksonville, FL [1 (link), 11 (link)]). A rabbit polyclonal antibody (ASYM24 07–414, MilliporeSigma, Burlington, MA) was used for immunohistochemistry for aDMA. Regions of interest were middle frontal gyrus (FCtx), motor cortex (MCtx) and hippocampus. Adjacent sections were immunostained for phospho-TDP-43 as above. All Immunoperoxidase sections were processed on a DAKO AutostainerPlus (Agilent/DAKO, Santa Clara, CA) with the DAKO EnVisionTM+ system-HRP with diaminobenzidine as the chromogen. Nonspecific antibody binding was blocked with normal goat serum (Sigma, St Louis, MO).
For double immunofluorescence staining, a rat monoclonal antibody was used to detect poly-GR (5H9, MilliporeSigma, Burlington, MA) and a rabbit polyclonal antibody was used to detect aDMA (MilliporeSigma, Burlington, MA). The fluorochromes were Alexa Fluor 568 and Alexa Fluor 488 conjugated to anti-rabbit or anti-rat IgG (Invitrogen/ThermoFisher, Waltham, MA).
For double immunofluorescence staining, a rat monoclonal antibody was used to detect poly-GR (5H9, MilliporeSigma, Burlington, MA) and a rabbit polyclonal antibody was used to detect aDMA (MilliporeSigma, Burlington, MA). The fluorochromes were Alexa Fluor 568 and Alexa Fluor 488 conjugated to anti-rabbit or anti-rat IgG (Invitrogen/ThermoFisher, Waltham, MA).
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