Control sections for ISH-IHC dual labeling in thick free-floating sections.

<p>A) Tissues were labeled with a rat positive control probe (i.e. rat peptidylprolyl isomerase B gene) provided by ACD. Clear ISH signal and IHC staining were observed. This probe acts as a positive control because it is present ubiquitously throughout rat tissue, albeit at modest levels in the cells depicted. B) Tissues were labeled with a negative control probe (i.e. <i>Bacillus subtilis</i> dihydrodipicolinate reductase gene) provided by ACD. Very infrequent ISH puncta are present, while strong IHC is clear. This probe was used as a negative control because it is a bacterial transcript not present in rat tissue. C) Tissue was labeled with Sprr1a ISH, but the primary TH antibody was excluded. As expected, strong Sprr1a ISH is apparent in the SN of the lesioned hemisphere (i.e. brown ISH puncta), but no TH IHC is present (i.e. lack of blue staining). D) To ensure the signals were dependent upon the Sprr1a ISH probe and TH primary antibody we omitted the Sprr1a probe and the TH primary antibody. As expected, this results in no signal for ISH (i.e. no brown puncta) or IHC (i.e. no blue staining). Scale bar: A-D = 50μm.