Fig. 2. Localization of trout cytoplasmic carbonic anhydrase (tCAc) protein in the gills of rainbow trout (Oncorhynchus mykiss) by immunohistochemistry. The images are overlays of three images collected individually for tCAc immunoreactivity (green), 5 immunoreactivity (red) and nuclei visualization (blue). Areas of overlap of tCAc and 5 immunoreactivity are indicated in yellow. Nuclei were visualized using 4',6'-diamidino-2-phenylindole. Medium (A) and high (D) magnification images representative of the sections examined indicate that cells displaying only tCAc immunoreactivity, only 5 immunoreactivity or co-localization of tCAc and 5 immunoreactivities were present in the lamellar epithelia as well as in the interlamellar regions. Omission of primary antibodies eliminated all immunofluorescence (B). Pre-absorption of the tCAc antibody using the peptide against which the antibody was raised eliminated tCAc immunoreactivity without affecting 5 immunoreactivity (C). Scale bars, 20 µm.