Light microscopic diagnosis of intestinal microsporidiosis is difficult with the use of routine histologic stains. This has led to an overreliance on transmission electron microscopic diagnosis. It was previously demonstrated that a modification of the standard Gomori one-step trichrome stain, using a 10-fold higher concentration of chromotrope-2R, can be used to detect microsporidial spores in stool. The use of the stain has now been extended to the detection of spores in sections of formaldehyde-fixed, paraffin-embedded intestinal biopsy specimens. Positive identification can be made of both intestinal species seen in patients with the acquired immunodeficiency syndrome, Enterocytozoon bieneusi and Septata intestinalis, when the diagnosis is inapparent or questionable on routine histologic analysis. The use of this simple stain should increase the sensitivity for diagnosing microsporidiosis by light microscopy, further obviating the need for transmission electron microscopy.