Fig. 5. A gradient of Math1 expression precedes the appearance of the early hair cell differentiation marker, MyosinVIIa. (A-D) Sections through the developing cochlear duct stained with antibody to Math1. (E,F) Sections stained with antibody to MyosinVIIa. (G,H) Sections double-labeled with antibodies to MyosinVIIa (red) and p27^Kip1 (green). (A,B) At E14.5, Math1 expression has spread to the medial region of the cochlear duct (A) from its site of initiation in the base (B, see Fig. 1 for reference). In the medial region (A), a single column of cells spanning the cochlear epithelium is seen, while in the base (B) two columns of Math1 stained cells are observed (arrowhead, medial; arrow, lateral). The Math1⁺ columns are separated by a row(s) of Math1^– cells. (C-H) One day later, at E15.5, the gradient of Math1 expression has progressed and two columns of Math1⁺ cells are seen in the apical region of the cochlear duct (C, arrowhead, medial; arrow, lateral), and Math1⁺ cells in the mid-basal region (D) have begun to resolve into the stereotyped pattern of one row of inner (arrowhead) and three rows of outer (bracket) hair cells. In contrast to the Math1 staining observed in the base (C,D), no MyosinVIIa staining is observed in adjacent sections in the apex (E), while only a single inner hair cell is stained with MyosinVIIa in the base (F, arrowhead), indicating that Math1 expression precedes that of MyosinVIIa. Sections double-labeled with antibody to p27^Kip1 (green) and MyosinVIIa (red) reveal the change from the thickened epithelium that is present in the undifferentiated apical region (G) compared with the bilayered epithelium in the differentiated organ of Corti (H). Scale bar: 50 µm.