Anti cdx2 mouse monoclonal antibody

To evaluate the quality of the cloned blastocysts from FD somatic cells, cell numbers were examined using immunofluorescence staining as previously described^{12 (link)}. The primary antibodies used were an anti-CDX2 mouse monoclonal antibody (1:500; BioGenex, San Ramon, CA, USA, MU392A-UC) for detecting the TE cells and an anti-Nanog rabbit polyclonal antibody (1:500; Abcam, Cambridge, UK, ab80892) for detecting the ICM cells. The secondary antibodies used were Alexa Fluor 488-labelled goat anti-mouse IgG (1:500; Molecular Probes Inc., Oregon, USA, A11029) and Alexa Fluor 568 goat anti-rabbit IgG (1:500; Abcam, Cambridge, UK, A11036). DNA was stained with DAPI (2 μg/mL; Molecular Probes).

Formalin-fixed, paraffin-embedded human colorectal tissues were stained with an anti-CDX2 mouse monoclonal antibody (Clone: CDX2-88, 1:50, BioGenex, San Ramon, CA, USA) using the avidin-biotin immunoperoxidase method. A scoring system described in a previous report [24 (link)] was used to quantify CDX2 expression levels. All IHC samples were evaluated independently by two researchers. The pathological therapeutic effects of a preoperative treatment are defined in the Japanese Classification of Colorectal Carcinoma, 8th Edition [39 ].

To count the total number of nuclei, as well as the number of trophectoderm (TE) cells and inner cell mass (ICM) blastomeres, immunofluorescence staining of blastocysts was performed. Blastocysts were fixed in PBS containing 1% paraformaldehyde for 1 h. The fixed embryos were washed twice in PBS containing 1% (w/v) Polyvinyl alcohol (PVA-PBS) for 15 min each and then stored in PVA-PBS containing 0.1% (v/v) Triton X-100 (35501–15, Nacalai Tesque, Kyoto, Japan) overnight at 4°C. The primary antibodies used were an anti-CDX2 mouse monoclonal antibody (1:500; BioGenex, San Ramon, CA, USA, MU392A-UC) to detect TE cells and an anti-Nanog rabbit polyclonal antibody (1:500; ab80892, Abcam, Cambridge, UK) to detect ICM cells. The secondary antibodies used were an Alexa Fluor 488-labelled goat anti-mouse IgG (1:500, A11001, Molecular Probes, Eugene, OR, USA) or Cy5-labeled goat anti-mouse IgG (1:500; ab97077, Abcam) and an Alexa Fluor 568-labelled goat anti-rabbit IgG (1:500; A11004, Molecular Probes). DNA was stained with 4′,6-diamidino-2-phenylindole (DAPI; 2 μg/mL; D1306, Molecular Probes). Total cell number was counted as DAPI positive cell.