Detection of apoptotic cells in cytology specimens: An application of TdT-mediated dUTP-biotin nick end labeling to cell smears

We applied TdT-mediated deoxyuridine triphosphate (dUTP)biotin nick end labeling (TUNEL) to cytologic smears in order to detect the cells undergoing apoptosis. These smears were obtained by scraping the cut surface of 9 cases of carcinoma, including renal-cell carcinoma (3 cases), esophageal squamous- cell carcinoma (3 cases), and gastric adenocarcinoma (3 cases), and were fixed and prepared by different methods. The results were also compared with those of tissue sections. TUNEL in smears was generally associated with higher background nuclear stain than in tissue sections. Smears that were fixed in 4% or 8% paraform aldehyde or absolute methanol exhibited results comparable with those of tissue sections, with minimum background in all cases examined. There were no significant differences in TUNEL labeling index among tissue sections and smears fixed in 4% or 8% paraformaldehyde or in absolute methanol. Smears treated in Carnoy's fixative (3:1 methanol:acetic acid) and air-dried smears demonstrated a higher background. TUNEL positivity could not be detected in slides decolorized from May-Gunwald-Giemsa stain. Markedly high background, which may occur as a result of artifactural DNA breaks, was also observed in slides decolorized from Papanicolaou stain, in which TUNEL-positive cells could be evaluated only in 3/8 cases. Application of the TUNEL method to cytology specimens has disadvantages or limitations compared to its application to histological sections, but the method is considered the most suitable one for detecting cells undergoing apoptosis in cytology materials.