The aim of this work is to evaluate the sensitivity and efficacy of a diagnostic approach based on scrapings of the superior tarsal conjunctiva examined by scanning electron microscopy (SEM). We examined 30 eyes of 26 consecutive patients with presumed microbial keratitis, who had no improvement after local broad-spectrum antibiotic therapy performed for at least 7 days at an effective dosage. 13 patients had negative culture tests for bacteria and fungi obtained from corneal curettage. After 72 hours of wash-out, scrapings of the upper tarsal conjunctiva for cytological examination of cell morphology by scanning electron microscope (SEM) were performed. With this method, the presence of pathogenic microorganisms was detected in 26/30 eyes and specific therapy was carried out on the basis of these results. Clinical improvement and eradication of organisms previously detected by SEM were observed in all positive patients within 1-7 weeks. No organisms were detected in four patients; in two of them the presence of inflammatory cells indicative of an allergic reaction (eosinophils and mast cells) was detected, in the other two there were signs of severe dry eye. The efficacy of this diagnostic technique is confirmed by the clinical response to treatment based on the microorganisms identified by SEM and by the eradication of previously detected microorganisms, verified by subsequent scanning electron microscopic (SEM) control examination. The results obtained support the validity of the proposed method and we believe that it constitutes a very useful tool for distinguishing, on the basis of the inflammatory infiltrate, the infectious forms from those of other nature and for detecting the presence of microorganisms resistant to the usual therapies.