DNA Labeling at Electron Microscopy

Abstract

Here, we describe a method for locating DNA on ultrathin sections. This technique is compatible with all usual fixation and embedding procedures and can be combined with cytochemical methods. Ultrathin sections are incubated in a medium containing terminal deoxynucleotidyl transferase (TdT) and various non-isotopic nucleotide analogs. The labeled nucleotides bound to the surface of ultrathin sections are then visualized by an indirect immunogold labeling technique. This high-resolution method provides a powerful tool for pinpointing the precise location of DNA within biological material, even where DNA is present in very low amounts.

Visualization of Chromatin in the Yeast Nucleus and Nucleolus Using Hyperosmotic Shock

“…Cells were fixed for 60 min at 4 • C in 1.6% glutaraldehyde in 0.1 M Sorensen's buffer (pH 7.4) and acetylated as previously described [40] or stained with the AgNOR technique [41]. After washing in Sorensen's buffer, cells were dehydrated through graded ethanol solutions and then processed for embedding in epon.…”