Immunohistochemical validation of calretinin expression in the 12th and SSp motor nuclei. (A) Darkfield coronal section of male zebra finch brainstem. (B) The same section as in A under fluorescence imaging shows calretinin expression (green) detected by a fluorescein isothiocyanate (FITC)-labeled calretinin-specific antibody. (C) Quantitative analysis of calretinin- expressing cells. The total number of motor neurons was determined from darkfield images; the number of calretinin-labeled neurons was determined from FITC fluorescence images. There are nine times as many calretinin-positive neurons in the SSp motor nucleus as there are in the 12th motor nucleus (n = 3; [*] p < 0.003). Scale bars, 500 μm.