Fig S1: Photographs of embryos E22 and E24. The bars in the photographs are 1 cm apart.

Fig S2: Visual representation and quantitative measures of contrast for unmodified T1-weighted, midsagittal magnetic resonance (MR) images (top left) and the corresponding contrast-enhanced MR images (top right) of embryos E28, E32, E35, E37, E39, E40, E42, E45, E50 upper and lower body, E65 distal fore- and hindlimb. Photographs of the embryos are shown (the bars in the embryo images are 1 cm apart) to allow comparison of the MR images with the embryo's gross morphology. In the histograms, red represents the unmodified sections and blue the contrast-enhanced sections. The histograms illustrate the results of histogram equalisation, namely a wider spread of intensity values, which in turn results in higher visual contrast, as can be seen by comparing the unmodified and contrast-enhanced MR images. The X-axis represents the possible grey values (scale: 0–255) and the Y-axis indicates the number of pixels found for each grey value.

Fig S3: Visual comparison of unmodified T1- and T2*-weighted midsagittal magnetic resonance imaging (MRI) sections of embryos E28, E32, E35, E37, E39, E40, E42, E45, E50 upper and lower body, E65 distal fore- and hindlimb (from left to right). As the embryos still largely consist of soft tissue, T2* weighting, which is sensitive to magnetic susceptibility interfaces, such as air–tissue or tissue–bone, provides less contrast in these images than T1 weighting, which reveals other magnetic resonance (MR) mechanisms, such as macromolecular structure, water compartmentalisation, mobility etc., that do vary between soft tissue types. The remaining trapped air bubbles distort the T2*-weighted images significantly. In the E65 images, the presence of bone marrow can be seen clearly, due to the contrast provided by its erythropoietic, iron-containing cells.

Fig S4: The three-dimensional (3D) reconstructions of embryos E39 and E45 are shown to give an impression of the advantage that 3D reconstructions offer to enhance understanding of embryogenesis.

Table S1: Overview of the imaging parameters used for each embryo. Due to the large variation in embryo size and tissue differentiation, the imaging parameters had to be adapted to each individual embryo. (TR = repetition time; TE = echo time; FOV = field of view).

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