Ab7759

Cultured cells were fixed in 4% PFA (Sigma-Aldrich) in PBS for 10 min and then permeabilized in PBS containing 0.3% Triton X-100 (Sigma-Aldrich). The cultures were then incubated with primary antibodies followed by secondary antibodies (see the dilutions below). 4′,6-diamidino-2-phenylindole (DAPI) (1:1,000) (Thermo Fisher Scientific) was added to visualize cell nuclei. Images were taken using the DMi8 inverted microscope (Leica). The primary antibodies used in the study were as follows: anti-HCG (ab9582, Abcam, 1:200), anti-dsRNA (MABE1134, Merck, 1:200), anti-ACE2 (ab15348, Abcam, 1:200), anti-GATA2 (WH0002624M1, Sigma-Aldrich, 1:100), anti-GATA3 (MA1-028, Invitrogen, 1:100), anti-SDC1 (12922, Cell Signaling Technology, 1:100), anti-DAB2 (ab76253, Abcam, 1:100), anti-MMP2 (40994, Cell Signaling Technology, 1:100), anti-SARS-CoV-2 nucleocapsid (MBS154642, MyBioSource, 1:300) and anti-HLA-G (ab7759, Abcam, 1:50). Secondary antibodies used in the study (all 1:400) were Alexa Fluor 488 goat anti-mouse IgG1 (A21121, Thermo Fisher Scientific), Alexa Fluor 555 goat anti-mouse IgG (A31570, Thermo Fisher Scientific), Alexa Fluor 555 goat anti-rabbit IgG (A21428, Thermo Fisher Scientific), Alexa Fluor 555 goat anti-mouse IgG2a (A21137, Thermo Fisher Scientific), Alexa Fluor 647 donkey anti-rabbit IgG (A31573, Thermo Fisher Scientific).

The villi tissue was frozen in liquid nitrogen and then sliced using a cryotome. The cryosections were incubated with rabbit anti-FTO (1:500, ab126605, Abcam), mouse anti-HLA-G (1:500, ab7759, Abcam) and mouse anti-m⁶A (1:500, ab208577, Abcam) antibodies, washed thrice with PBS, and then probed with the secondary antibody for 1h. After washing thrice with PBS, the sections were sealed with the coverslips and gelvatol, and observed under a fluorescence microscope (Leica TCS-SP5).