Fluorescence Microscopy in Exfoliative Cytology

Abstract

EXFOLIATIVE cytology is a diagnostic procedure, which, having been generally accepted, is growing rapidly in importance as a means of early diagnosis of cancer. The Papanicolaou technique, developed originally to investigate gynaecological cancer, has been extensively applied to material from various sources and hag collected a voluminous literature testifying to its usefulness, (Papanicolaou, 1954;Ayre, 1951; Schade, 1959). The diagnosis of the malignant cell however is, in the final analysis, subjective and it is experience and familiarity with a technique that is important, not the stain used. Philps (1954), using temporary wet films stained with methylene blue (Schuster, 1947) and permanent films stained with haemalum and eosin (Dudgeon and Wrigley, 1935), diagnosed 76-9 per cent carcinomas of lung by means of exfoliative cytology. Spriggs (1957) recommends May-Grundwald-Giemsa for the examination of serious fluids. Luckeock (1961), reporting from the Brompton Hospital, describes the use of Ehrlich's acid haematoxylin and eosin for the examination of tumour cells in blood, whereas, for the same purpose, Alexander and Spriggs (1960) find Romanowsky stains useful. The present study was undertaken to evaluate fluorescence microscopy for cytological diagnosis using acridine orange as described by von Bertalanffy, Masin and Masin (1958).Fluorescence microscopy has been applied to cytology using berberine sulphate, acid fuchsin and the acridine group of stains (Friedman, 1950; Mellors, Glassman and Papanicolaou, 1952). Its use is based on the affinity of the basic fluorochrome dyes for the nucleic acids. Studies of the neoplastic cell have shown an increase in the total nucleic acids and ribonucleic acid (RNA). This increase is characteristic of cells showing a high protein synthesis and is seen in embryonic, regenerating, secretory and malignant cells. The RNA is found in the cytoplasm and the nucleoli of the affected cells. Increase in deoxyribonucleic acid (DNA), found in the chromatin of the nucleus, is seen in cells undergoing mitosis and polyploidy characteristic of advanced cancer. The fluorescent techniques described by Friedman and others have proved tedious and inadequate.von Bertalanffy et al. (1958) have reviewed the literature on the role of nucleic acids in malignant cells, and the theoretical considerations for the use of basic fluorochrome stains and they have suggested the use of acridine orange in routine exfoliative cytology.Acridine orange (AO) is a histochemical fluorochrome with a selective affinity for nucleic acids. At a concentration of 0.01 per cent and a pH of 6, recommended by von Bertalanffy, the DNA fluoresces yellow to whitish green, and the RNA red.

Confocal microscopy and exfoliative cytology

“…Exfoliative cytology is a diagnostic procedure which has been generally accepted, is growing rapidly in importance as a means of early diagnosis of cancer. [ 6 ] The diagnosis of malignant cell using cytological smears, however, is in the final analysis, subjective and it is experience and familiarity with a technique that is important, not the stain used.…”

“…In addition, presence of irregular nuclei and mitotic figures in the smears add to the criteria for malignancy. [ 6 13 14 ]…”

Comparative cytopathologic evaluation using acridine orange with Papanicolaou stain in psychoactive substances abusers with potentially malignant and malignant disorders

“…Exfoliative cytology is a noninvasive diagnostic procedure, which is growing rapidly in importance as a tool of early diagnosis of cancer. [ 8 ] Early detection of PS abusers with potentially malignant lesions and malignant lesions in the oral cavity could be greatly improved through techniques that permit visualization of subtle cellular changes indicative of the neoplastic transformation process. One such technique is fluorescence microscopy.…”

“…The application of fluorescence microscopy results in increased brilliance depending on the various degrees of polymerization of nucleic acid in malignant cells. [ 6 8 ]…”

The exfoliative cytology of endometrial cancer with fluorescence microscopy.

“…Cytological examination of body fluids other than vaginal and cervical discharge carried out at the Royal Free Hospital had shown that the technique is inferior to the Papanicolaou method for the detection of squamous carcinoma, especially a well differentiated one, exfoliating keratinized malignant cells, but offers considerable advantage in the diagnosis of adenocarcinoma and anaplastic tumours, and helps to differentiate between cells of columnar origin and histiocytes (Grubb and Crabbe, 1961). The few references in the literature to the red and yellow fluorescing endometrial cells (Hamperl and Schummelfeder, 1961;Elevitch and Brunson, 1961;Marks and Goodwin, 1962;Sani, Citti and Caramazza, 1962) appeared to suggest that endometrial cells may be more easily discerned in cytological smears stained with acridine orange than is the case with Papaninolaou smears.…”