BACKGROUND: American cutaneous leishmaniasis is a disease with a wide variety of clinical manifestations that is expanding throughout Brazil, the state of Mato Grosso do Sul constituting a significant endemic area. OBJECTIVES: To evaluate the clinical, epidemiological and laboratory characteristics of patients with American cutaneous leishmaniasis. Patients were recruited among those attending the Maria Aparecida Pedrossian Teaching Hospital of the Federal University of Mato Grosso do Sul, Brazil. METHODS: This was a cross-sectional, observational study conducted using a descriptive and analytical approach. Data from patients suspected of having American cutaneous leishmaniasis who were receiving care at this institute between 1998 and 2008 and were referred to the institute's parasitology laboratory for confirmation of diagnosis were evaluated retrospectively. Clinical and laboratory criteria were taken into consideration for the inclusion of patients to the study. RESULTS: Forty-seven patients were included in the study, the majority of whom were male and between 45 and 59 years of age. Most had the cutaneous form of the disease with a single, ulcerated lesion on exposed areas of the body, which had generally been present for periods of less than six months. Mucosal involvement increased with age and was highest in patients who had sought medical care at a later stage. The Montenegro skin test showed the highest sensitivity. Finding the parasite was more difficult in older lesions. CONCLUSION: Suspicion of the disease at an early stage is of extreme importance for a precise diagnosis. A combination of parasitological and immunological tests renders laboratory diagnosis more reliable.