DETECTION OF METHYLCYTOSINE-RICH HETEROCHROMATIN ON BANDED CHROMOSOMES - APPLICATION TO CELLS WITH VARIOUS STATUS OF DNA METHYLATION

A procedure including incorporation of 5-bromodeoxyuridine (BrdU) in D NA and a thermal denaturation step was developed to obtain both R-band ing and efficient binding of anti-5-methylcytosine antibodies on metap hase chromosomes. BrdU incorporation improved the efficiency of antibo dy binding disclosed by immunofluorescence staining. This method allow ed semiquantitative analysis of the antibody binding sites on straight forward characterized metaphase chromosomes and was applied to normal human lymphocytes and lymphoblastoid cell lines for which DNA methylat ion status had been previously analyzed. A correlation was established between level of DNA methylation and the semiquantitative estimate of antibody fixation. This procedure can be used to study DNA methylatio n on metaphase chromosomes in transformed and cancerous cell lines.