Hoechst 33258

First-trimester placental tissue (FiTPT) at 7–11 weeks of gestation (n=7) and full-term placental tissue (FTPT, n=15) were fixed in 4% PFA for 24 h, embedded in paraffin, and sectioned according to standard histology methods. Paraffin-embedded tissues were deparaffinized in 2 changes of xylene and rehydrated in decreasing concentrations of ethanol.
Antigen retrieval was performed in Tris-EDTA buffer (10 mM Tris Base, 1 mM EDTA solution, pH 9.0) at +95°С for 30 min. Nonspecific reactivity was reduced by incubating tissue sections in blocking solution (0.5% BSA and 1% goat serum in 0.1 M PBS with 0.3% Triton X-100) for 30 min.
Immunostaining was accomplished by overnight incubation at +4°С with primary antibodies (Table S3) diluted in Antibody Diluent (Dako, Denmark). After extensive washing with 0.1 M PBS, samples were incubated in the appropriate species-specific secondary antibodies (Table S3) diluted 1:1000 in Antibody Diluent (Dako, Denmark) for 1 h at RT in the dark. After nuclear staining with Hoechst 33258 (Vector Laboratories, UK), slides were mounted in Mowiol® 4-88 (Sigma, USA).
Confocal analysis was performed with a Zeiss LSM 510 Meta microscope (Zeiss, Germany) and images were captured with Zeiss LSM Image Browser software.

T47D and MCF7 tumor xenografts from NOD/SCID mice were fixed in 10% phosphate-buffered formalin and embedded in paraffin. Paraffin sections were cut at a thickness of 2 μm, mounted on SuperFrostTM Plus microscopy slides (Menzel-Gläser), and dried at 70 °C for 2 h. The sections were dewaxed in xylene and rehydrated according to the standard histopathological procedures. The slides were first placed in the solution of Tris-EDTA pH 9.0 and autoclaved for 7.5 min for antigen retrieval. The slides were then blocked with 3% hydrogen peroxide for 15 min and incubated with biotin conjugated UEA-1 lectin (Vector Laboratories) for 1 h at room temperature followed by fluorescein-conjugated streptavidin (Jackson ImmunoResearch) for 1 h at room temperature. Finally, the slides were stained with Hoechst 33258 for 15 min and mounted with Vectashield (Vector Laboratories). For Globo H staining, the slides were stained using Benchmark XT machine (Ventana). The slides were added with anti-Globo H antibody (Vk9) in power block HK085-5K solution (Ventana) at 37 °C for 1 h followed by biotin conjugated anti-IgG and streptavidin-HRP for 20 min and applied with iVIEW DAB Detection kit (Ventana). Leica autostainer XL (CV5030) machine was then used for counter staining with Mayer’s hematoxylin and mounting. The images were collected with Leica automatic upright microscope (DM 6000B).