Extended Data Fig. 1: Analysis of the pHi in the chicken embryo in vivo.
From: Intracellular pH controls WNT downstream of glycolysis in amniote embryos
a, Ratiometric live expression of pHluorin (488/405 nm) detected in the posterior domain of electroporated embryos exposed to different pH buffers and nigericin and valinomycin (n = 7). Fluorescence intensity is shown by a pseudocolour image (Fire colour) using ImageJ. The yellow signal indicates lower pH. The ventral view shows the anterior region to the left. Scale bars, 100 μm. b, Each dot represents the average 488/405-nm signal ratio of about 300 single cells segmented in one embryo (n = 2 for pH 5.5, n = 3 for pH 6.5 and n = 2 for pH 7.5). Embryos were treated for 20 min in different pH buffers with the protonophores nigericin and valinomycin, before live imaging. The red horizontal bar is the mean and error bar is the s.d. c, Micro-dissected posterior PSM incubated with 20 μM BCECF (n = 7). Fluorescence intensities for excitation at 405 nm (blue) and 488 nm (green) are shown (left). In addition, the 488/405-nm ratio is shown (right). Scale bars, 100 μm. A, anterior; P, posterior. d, Fluorescence 488/405-nm ratios along the PSM. Each coloured line corresponds to an explant (n = 7 from two independent experiments). med, medial PSM; pos, posterior region. Two-sided, paired t-test. **P = 0.009. e, f, Whole-mount in situ hybridization of 2-day-old chicken embryos cultured at different pH and hybridized with MSGN1 (pH 5.3: n = 4, pH 7.2: n = 3 and pH 7.6: n = 3) (e) and SAX1 (pH 5.3: n = 6, pH 7.2: n = 4 and pH 7.6: n = 5) (f). The ventral view shows the anterior region to the top. Scale bars, 100 μm.
