Table 2.
Antibodies specifications and working solutions used in this study.
| Target | m/pAb | Specie | Source | Cat. Number | Dilution | Assay |
|---|---|---|---|---|---|---|
| LIMD2 1 | pAb | Rabbit | Peng et al. [8] | - | 1: 1000 | WB |
| LIMD2 2 | pAb | Rabbit | Abcam | Ab221110 | 1:33 | WB, HCS |
| Α-tubulin | mAb | Mouse | Sigma-Aldrich | T9026 | 1:10,000 | WB |
| F-actin 3 | - | - | Thermo Fisher Scientific | A12379 | 1:300 | HCS |
| Histone-γH2AX | pAb | Rabbit | Novus Biologicals | NB100-384 | 1:100 | FC |
| P53 | pAb | Rabbit | Abcam | ab131442 | 1:100 | FC |
| β-catenin | mAb | Mouse | Santa Cruz Biotechnology | Sc-376841 | 1:100 | FC, HCS |
| STAT3 (Y705) | pAb | Rabbit | R&D Systems | AF4607 | 1:100 | FC, HCS |
| STAT5α | mAb | Mouse | Santa Cruz Biotechnology | Sc-166479 | 1:200 | FC, HCS |
| SLUG | pAb | Rabbit | Santa Cruz Biotechnology | sc-15391 | 1:100 | FC, HCS |
| TWIST2 | mAb | Mouse | Abcam | Ab50887 | 1:50 | FC, HCS |
| N-cadherin | mAb | Mouse | Dako | M3613 | 1:100 | FC, HCS |
| FITC-conjugated goat anti-rabbit IgG | pAb | Goat | Thermo Fisher Scientific | AB42978 | 1:1000 | FC, HCS |
| FITC-conjugated goat anti-mouse IgG | pAb | Goat | Thermo Fisher Scientific | AB2533946 | 1:10,000 | FC, HCS |
WB, Western blot; FC, flow cytometry; HCS, high-content screening. 1 LIMD2 antibody developed by Peng et al. [8] was used to screen the LIMD2 expression in cell lines derived from different histological types of thyroid cancer. This antibody is now commercialized by Merck (MABC1168) and recognizes the full-length human LIMD2. 2 LIMD2 antibody Ab221110 was used to confirm the LIMD2 knockout in both edited cells, since it recognized the amino acid residues 105–126 (C-terminal region) comprised within the edited region. 3 F-actin was detected using the FITC-conjugated phalloidin mycotoxin.