Pregnant Cmas^+/– mice were treated at E4.5 and E6.5 either with PBS (n = 3) or CVF (n = 3) to deplete maternal C3. E8.5 sagittal uteri paraffin sections (A and B). (A) The reduced size of the EPC and lack of a CP in Cmas^–/– embryos of PBS-treated mothers was reverted to the control phenotype upon CVF treatment, as visualized by immunohistochemical cytokeratin-8 staining. The lack of CEBPB reactivity (indirect immunofluorescence) in Cmas^–/– embryos of PBS-treated mothers was restored upon CVF treatment. PNA reactivity documenting the loss of cell surface sialylation was maintained in Cmas^–/– embryos of PBS- and CVF-treated mothers, indicating that the asialo phenotype was not influenced by CVF. Representative images of experiments with PBS-treated mice: control, n = 5; Cmas^–/–, n = 3 embryos; CVF treated mice: control, n = 5; Cmas^–/–, n = 4 embryos. (B) Collagen IV indirect immunofluorescence (red). Thickened RM (arrow) in Cmas^–/– embryos of PBS-treated mothers was converted to the control phenotype upon CVF treatment. Parietal endoderm is marked by arrowheads. Nuclei shown in white were stained with DAPI. (C) Quantification of RM thickness measured on collagen IV immunofluorescence images at the anti-mesometrial pole (PBS: control, n = 6; Cmas^–/–, n = 3 embryos; CVF: control, n = 5; Cmas^–/–, n = 4 embryos). (D) Mean of fetal size as measured by the sum of the areas of the amniotic cavity, exocoelomic cavity, ectoplacental cavity, and embryo proper in (μm²/10⁵) (PBS: control, n = 5; Cmas^–/–, n = 4 embryos; CVF: control, n = 5; Cmas^–/–, n = 3 embryos); a schematic of the areas is shown in Supplemental Figure 5. Statistical analyses by 1-way ANOVA with Newman-Keuls post test (*P < 0.05; **P < 0.01; ***P < 0.001). Error bars indicate SD (C and D).