Trichophytia profunda acuta of the glabrous skin (TPAGS) arose in a 67-year-old Japanese man. The patient presented indurated erythematous plaques and nodules on his left forearm. Direct microscopic examination of the scale in KOH preparation was negative for fungal elements, and culture for dermatophytes was also negative. Although fungal infection could not be proven in hematoxillin-eosin stained sections, deep-cut sections of the biopsied skin lesion with PAS stain revealed the ectothrix presence of fungal elements. Nested PCR was done with Trichophyton specific primers directed to internal transcribed spacer gene 1 (ITS1), using template DNA obtained from formalin fixed, paraffin embedded skin sections. A single band corresponding to T. rubrum was obtained, and the etiological agent was thus identified. KOH tests and cultures may often turn out unsuccessful, perhaps reflecting the hair follicle dominant fungus growth in TPAGS. Although these tests are most important for diagnosis of TPAGS, nested PCR using paraffin embedded skin sections may be an alternative method to identify the etiological agent.