Fig. 2. Analysis of progenitor cell and myocardial markers in explants after 12 and 72 hours of culture. (A-F) Posterior (p) SHF explants at the 4- to 6-somite stages after 12 (A-C) and 72 (D-F) hours of culture. (A,D) Hoechst staining; (B,E) Islet1 (Isl1) immunofluorescence; (C,F) myosin heavy chain (MHC) immunofluorescence. The analysis has been repeated on five explants for each time point. (G) RT-PCR analysis on right (R) and left (L) explants (five pooled explants for each side) at the 4- to 6-somite stage after 72 hours of culture, with primers for Mlc2a, COUP-TFII (Nr2f2) and Mlc2v mRNAs. cDNA from the common atrium (A) or from the ventricular region (V) of five hearts from E9 embryos. Controls (-) contained RNA without reverse transcription. β-actin transcripts indicate similar quantities of RNA. (H-K) An explant of the pSHF from Mlc1v-nlacZ-24 embryos at the 4- to 6-somite stage after 72 hours of culture. (H) Hoechst staining; (I) MHC immunofluorescence; (J) β-gal immunofluorescence; (K) merge of H,I,J. Scale bars: 10 µm in F for A-F; 30 µm in H-K.