Anti-5-methylcytosine (5-mC) rabbit monoclonal antibody [RM231]

ELISA of single stranded DNA using rabbit monoclonal anti-5-mC (clone RM231) antibody. The plate was coated with streptavidin and then biotinylated single stranded unmethylated DNA, 5-Methylcytosine (5-mC) DNA, and 5-Hydroxymethylcytosine (5-hmC) DNA. A serial dilution of RM231 was used as the primary antibody, and an alkaline phosphatase conjugated anti-rabbit IgG as the secondary antibody.

Dot blot of double stranded DNA using rabbit monoclonal anti-5-mC (clone RM231) antibody. The membrane was pre-spotted with 50, 5, and 0.5 ng/dot of double stranded 5-Hydroxymethylcytosine (5-hmC) DNA, 5-Methylcytosine (5-mC) DNA, and unmethylated DNA. The pre-spotted membrane was then blotted with RM231.

Immunocytochemical staining of HeLa cells using rabbit monoclonal anti-5-mC (clone RM231) antibody (red). Actin filaments have been labeled with fluorescein phalloidin (green), and nuclei stained with DAPI (blue).

Direct ELISA of HeLa cell genomic DNA using anti-5-mC antibody (RM231). The plate was directly coated with different concentrations of genomic DNA isolated from HeLa cells. 1 µg/mL or 3 µg/mL of RM231 was used as the primary antibody, and a HRP conjugated anti-rabbit IgG as the secondary antibody.

MeDIP was performed using anti-5-mC antibody (RM231) at a 2:1 DNA: Ab ratio. 1 ng of unmethylated, 5-Methylcytosine (5-mC) or 5-Hydroxymethylcytosine (5-hmC) DNA standard (897 bp) was spiked in 1 µg of genomic DNA isolated from HeLa cells as the control. Realtime PCR was then performed to determine the capture of DNA standard as in % of input.

Flow Cytometry analysis of 5-mC expression in HEK293 cells using Anti-5-mC rabbit monoclonal antibody, Clone RM231. The cells were fixed with ice-cold MeOH, permeabilized with 0.5% Triton X-100, denatured with 2N HCl, then stained with RM231 (anti-5-mC, Blue) or with a negative control antibody, Clone RM105, Red.