Comparison of Western blot and mass spectrometry analysis. We probed the synaptosomal fraction with different antibodies to confirm our quantitative mass spectrometry results. Immunoblot analysis was performed twice on three different synaptosomal fractions from p1 and p45, and the intensity of the immunoreactivity was measured. A representative immunoblot is shown for each antibody tested, and a bar graph with the immunoreactivity measurements from the synaptosomal preparations (black). N = 6 for each development period for each antibody. The Y-axis represents mean pixel intensity. Proteins that demonstrated a significant change in immunoreactivity in the synaptosomal fraction were also tested to determine if the significant change was also presented in total cerebellar homogenate (white). Statistical significance was determined by a two-tailed t-test. The following antibodies were employed: (A) Glutamate receptor subunit 2/3, (B) Na+/K+ ATPase alpha-1 chain, (C) alpha-tubulin, (D) cytochrome c oxidase subunit 1. Next to the immunoblot is the SILAM analysis of the same protein. The white bars represent the p1 sample, and the black bars represent the p45 sample. The Y-axis represents the 14N/15N ratio. The SILAM data were determined to be significantly altered (P < 0.05) during post-natal development by one-way ANOVA analysis followed by a Bonferroni post-hoc test to determine the significant difference of the protein expression between p1 and p45. The SILAM bar graph for alpha-tubulin represents the average mass spectrometry data for four proteins: tubulin alpha-1, tubulin alpha-2, tubulin alpha-3, and tubulin alpha-8. *P < 0.01; **P < 0.001.
