Enhanced numbers of progenitors and arrest of neutrophil differentiation in Pten mutants. A panel of in situ hybridization markers for blood lineages was used on 5 dpf wild-type and Pten mutant (ptena^−/−ptenb^−/−) embryos: (A-B) Globin and (C-D) gata1, both indicative of erythroblasts; (E-F) l-plastin, marking lymphocytes, macrophages, and neutrophils; (G-H) mpo, indicating neutrophils; (I-J) c-fms, indicative of macrophages. (K-N) At 5 dpf, SB staining was used to assess the number of mature neutrophils in various parts of the embryo. Panels K′-N′ represent magnifications of the boxed areas in the corresponding panels. (O) SB⁺ cells were counted in the head and yolk sac (mainly the primitive wave) and in the trunk, tail, and CHT region (mainly the definitive wave) as indicated in supplemental Figure 2. The results are expressed as average number of cells per embryo with the error bars indicating SEM; wild type, n = 6, ptena^−/−ptenb^−/− (n = 7). Normal distribution of data points was assessed with the Shapiro-Wilk test. Statistical comparison of groups was performed by the 2-tailed t test. *P < .05, **P < .01, ***P < .001 as indicated. Representative embryos are depicted; scale bars in panel J (200 µm) and N (100 µm) are representative for panels A-J, respectively, panels K-N. wt, wild type; mt, Pten mutant.