Methyl Green Staining and DNA Strands In Vitro: High Affinity of Methyl Green Dye to Cytosine and Guanine

Abstract

We occasionally encounter decreased staining of methyl green (MG) in heated sections for antigen retrieval. To examine the relation between MG staining and conditions of DNA strands, DNA in various conditions were blotted on the transfer membrane and stained. It was found that staining intensity of MG was changed due to structural alterations of DNA strands. Double stranded DNA in various solutions, denatured single-stranded DNA, DNase treated samples, and short length single-stranded oligonucleotides were examined. DNA diluted in high concentrated SSC was stained more strongly than low concentrated SSC or distilled water. Denatured and/or DNase treated DNA showed decreased staining. It was especially noteworthy that oligonucleotide DNA strands of poly C and poly G strands were positively stained, but very faint MG staining was also observed for poly A while no staining was noted for poly T. In conclusion, we presented that MG staining intensity was altered by the conditions of DNA in vitro. Intense MG staining for basic acids with three hydrogen bonds, cytosine and guanine, suggest a possible mechanism for MG staining.