Determination of DNA Methylation and Hydroxymethylation Levels in Biological Samples by Field-Amplified Sample Injection-Capillary Zone Electrophoresis with UV Detection

摘要

In this study, a method of field-amplified sample injection coupled with capillary zone electrophoresis with ultraviolet detection was established for evaluation of DNA methylation and hydroxymethylation levels in biological materials. By modifying an existing method, the separation of cytosine (C), 5-methylcytosine (5-mC) and 5-hydroxymethylcytosine (5-hmC) was performed on an uncoated capillary column (40 cm x 75 mu m I.D.) using 300 mmol L-1 tris solution (pH 2.90) as running buffer and detected at 280 nm. The detection limits (S/N = 3) of the method were 0.004 ng mL(-1) for cytosine (C), 0.01 ng mL(-1) for 5-methylcytosine (5-mC), and 0.02 ng mL(-1) for 5-hydroxymethylcytosine (5-hmC). The proposed method has been successfully applied to the evaluation of DNA methylation and hydroxymethylation levels of blood samples from 15 hepatocellular carcinoma patients and 5 liver cirrhosis patients and liver tissues from 50 pairs of tumor and matched tumor-adjacent samples.