This is a knockout-validated antibody summary, based on publications "N 6-methyladenosine of Chromosome-Associated Regulatory RNA Regulates Chromatin State and Transcription", as cited below [1], "RNA m6A methylation orchestrates cancer growth and metastasis via macrophage reprogramming" for western blot knockout validation (figure 4d, 4i, s1e) [2], and "The essential roles of m6A RNA modification to stimulate ENO1-dependent glycolysis and tumorigenesis in lung adenocarcinoma" for western blot knockout validation (figure s2b,s2c) [3]. Labome curates formal publications to compile a list of antibodies with unambiguous specificity within Validated Antibody Database (VAD).

Rabbit monoclonal IgG

Company: Abcam

Antibody: METTL3

Catalog number: ab195352

Summary: Rabbit monoclonal IgG against a recombinant fragment within human METTL3 (aa 1-250). Reacts with mouse, rat, human and pig. It is predicted to work also with sheep, goat, horse, guinea pig, cow, cat, dog, non-human primates. Suitable for western blot, immunoprecipitation, immunocytochemistry/immunofluorescence, immunohistochemistry (paraffin and frozen), and flow cytometry.

Western blot

Control and Mettl3 KO mESCs. Samples were homogenized in RIPA buffer (Invitrogen) containing 1 × protease inhibitor

cocktail and 1 × phosphatase inhibitor cocktail (Sigma).

5% milk PBST for 30 min at room temperature.

Diluted primary antibody solution at 4 °C overnight.

A dilution of secondary antibody conjugated to HRP for 1 h at RT.

SuperSignal West Dura Extended Duration Substrate kit (Thermo) and FluroChem R (Proteinsimple).

Immunoblot assays showing METTL3 protein level in Mettl3 KO and rescue with WT and inactive mutant METTL3. mESCs compared with the controls. The protein levels were quantified by normalized to GAPDH protein level. EV, empty vector. Please see Figure S1a in the article [1].