MethylFlash Methylated DNA 5-mC Quantification Kit

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	Do not allow pipette tip to touch the outer edges or inner sides of the wells to prevent solutions from sticking to the surface

Protocol tips
Do not allow pipette tip to touch the outer edges or inner sides of the wells to prevent solutions from sticking to the surface

Publication protocol

Global DNA methylation
DNA was extracted from cultured cells by standard methods (DNeasy system, Qiagen). For the measurement of global DNA methylation levels, 5-methyl-2′-deoxycytidine (5mdC) and 2′-deoxyguanosine (dG) were determined by an HPLC-based method.84 Normalized 5mdC levels were calculated as percentage 5mdC/dG. When indicated, global DNA methylation was measured by the MethylFlash system (Epigentek). The assay yielded 5mdC values as percentage of PicoGreen-quantified input DNA (wt/wt). Each value was multiplied by 4 to obtain a final value comparable to that obtained by the HPLC-based method. The MethylFlash assay yielded responses to AA and OA in THP-1 monocytes, that were consistent with the ones obtained by the HPLC-based 5mdC quantitation, except for baseline (unstimulated cells) values that were 5% lower (P>0 0.05, data corresponding to a total of 15 and 21 experiments, respectively).

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Manufacturer protocol

Download the product protocol from Epigentek for MethylFlash Methylated DNA 5-mC Quantification Kit below.

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