Fig. 5. Blocking the cardiac cell cycle results in loss of early, but not late, cardiac markers. (A) Cardiac explants isolated and cultured in DMSO or aphidicolin to block cells in S phase beginning at stage 22 and fixed at stage 37. Whole-mount immunostaining of explants with phosphohistone H3-specific antibody (red). In situ hybridization on explants with the early cardiac markers (B) Tbx5, Tbx20, Nkx2.5, (C) Gata4, Gata5 and Gata6, (D) with the cardiac differentiation markers Hsp27 and MLC1v', and (E) by whole-mount immunostaining with the cardiac differentiation markers Tmy and MHC. (F) Explants were treated with DMSO or colchicine (Colch) to block cells in M phase of the cell cycle. In situ hybridization was performed to examine expression of Tbx5, Nkx2.5 and MLC1V', as indicated. Aph, aphidicolin; BF, bright field; pH3, phosphohistone H3-specific antibody.