Summary Patients receiving high-dose chemotherapy (HD-CT) are at risk of severe mucositis. Most prevention studies evaluate the degree of mucositis on clinical, and therefore subjective, measurements. The aim of this study was to develop an objective in vitro assay of chemotherapy-induced mucositis. Twelve patients with locally advanced breast carcinoma received HD-CT followed by peripheral stem cell reinfusion. Before and twice weekly after HD-CT, the mucosa was evaluated by an oral washing, a buccal smear and the World Health Organization (WHO) toxicity grading; furthermore, blood leucocyte levels were determined. For the oral washings, the percentage of viable epithelial cells was determined by trypan blue dye exclusion and leucocytes were counted by fluorescence microscopy after incubation with acridine orange. Maturity of buccal cells was assessed by staining buccal smears for morphology according to Papanicolaou (Whitacker D and Williams V, 1994). Eight healthy volunteers served as controls. The mean percentage (± s.e.m.) of viable oral epithelial cells was stable in controls (44 ± 2%). In patients, they increased after HD-CT, which was significant after day 7 compared with pretreatment (P < 0.05). In addition, a shift from mature to immature epithelial cells in buccal smears was observed. Oral leucocyte levels were closely correlated with the blood leucocyte counts. The WHO score followed the results of these other evaluations with some delay. The viability of buccal cells obtained by oral washings increases after HD-CT. This is possibly because of desquamation of the upper oral mucosa layer, with a shift from mature to more immature cells. These data can be quantitated, and this assay may therefore be useful in studies aimed at prevention of mucositis.Keywords: mucositis; in vitro assay; quantitation Mucositis is a common, always unpleasant, sometimes unbearable toxic side-effect of chemotherapy. In particular, in patients receiving high-dose chemotherapy followed by bone marrow or peripheral stem cell transplantation, mucositis can be dose limiting. Chemotherapy causes a direct toxic effect on the rapidly dividing cells of the basal oral epithelium, which can result in mucosal atrophy, erythema and ulceration. The severe stages of mucositis with disruption of the oral mucosal barrier can lead to mucosal ulcers and secondary infection. In addition, it can provide a portal of entry for micro-organisms into the systemic circulation, which can lead to life-threatening septicaemia in myelosuppressed patients. Mucositis causes major discomfort, such as pain requiring intensive analgesia, and may restrict or even prohibit normal oral feeding and drug intake (Sonis, 1989;Sonis et al, 1990;Toth et al, 1990;Peterson, 1992; Woo et al, 1993).Grading of mucositis is necessary to document its degree and to evaluate the effect of measures for prevention or intervention. Most available scoring systems are based on a combination of objective changes in the mucosa (e.g. erythema, ulceration), subjective complaints (e.g....