Extended Data Fig. 4: Validation of DCX⁺ signal specificity.

a, Four distinct anti-DCX antibodies were used. The company, host species, RRID, immunogen, concentration of use, and signal enhancement protocol applied are indicated. b, Schematic diagram of the DCX protein. The anti-DCX antibodies used were raised against different domains of the protein. c, Number of DCX⁺ cells detected by using the distinct anti-DCX antibodies, with or without NaBH₄ and HC-AR sample pretreatment (one-way ANOVA, F_8,32 = 15.07, P < 0.0001). The graph represents mean values ± s.e.m. NaBH₄ + HC-AR incubation was required to adequately detect DCX⁺ cells for all the antibodies tested, although polyclonal goat anti-DCX (Santa Cruz) and mouse anti-DCX (Santa Cruz) antibodies gave the best results. d, Percentage of DCX⁺ cells double labeled by a polyclonal goat anti-DCX antibody (Santa Cruz) and other anti-DCX antibodies (one-way ANOVA, F_6,24 = 27.05, P < 0.0001). The graph represents mean values ± s.e.m. e, Percentage of DCX⁺ cells double labeled by a monoclonal mouse anti-DCX antibody (Santa Cruz) and other antibodies (one-way ANOVA, F_6,24 = 9.754, P < 0.0001). The graph represents mean values ± s.e.m. f,g, Representative images showing double labeling of DCX⁺ cells by using a polyclonal goat anti-DCX antibody (Santa Cruz) and a monoclonal mouse anti-DCX antibody (Santa Cruz) (f) or a polyclonal goat anti-DCX antibody (Santa Cruz) and a polyclonal rabbit anti-DCX antibody (Atlas-Sigma) (g). On the basis of signal specificity, signal/background ratio, tissue penetration, and working concentration, we opted to use the polyclonal goat anti-DCX antibody (Santa Cruz) to perform all subsequent quantifications. h, Representative image of a DCX⁺ cell counterstained with Nissl and visualized by immunohistochemistry. i, Dot-blot experiment showing no signal after preadsorption of polyclonal goat anti-DCX antibody (Santa Cruz) with a control synthetic DCX peptide that comprised the C-terminal domain of the protein. The full-length unprocessed blot is shown as source data. j, Representative image of DCX staining after sample incubation with preadsorbed polyclonal goat anti-DCX antibody (in red) showing no signal in the red channel. In c–h, n = 5 control subjects. Twenty measurements were performed for each subject. In i and j, the experiment was repeated twice with identical results. Yellow scale bars, 50 μm; blue scale bars, 10 μm; black scale bars, 20 μm. Magenta triangles indicate DCX⁺ cells. *0.05 > P ≥ 0.01; **0.01 > P ≥ 0.001; ***P < 0.001. In c–e, asterisks indicate changes with respect to the samples that did not receive NaBH₄ + HC-AR treatment in Tukey post hoc comparisons.

Source Data