Development and Validation of Ultra-Rapid Periodic Acid-Schiff Stain for Use in Identifying Fungus on Frozen Section

There is no standardized process for utilization of periodic acid-Schiff during intraoperative frozen sections to identify fungal organisms.

To develop a rapid staining process for fresh tissue with periodic acid-Schiff during intraoperative consultation and develop an appropriate control block.

Muscle tissue was inoculated with 2 species of fungus (Aspergillus fumigatus and Paecilomyces spp) and grown at 3 different temperatures for 72 hours. Inoculated tissue was embedded in optimal cutting temperature compound, cut, and stained using a modified periodic acid-Schiff stain. The optimal control was determined for future use as the standard control. Multiple control slides were cut and stained, using successively shorter time intervals for each step. The staining process that provided accurate results in the shortest amount of time was deemed ultra-rapid periodic acid-Schiff. This method was validated by carryover studies and clinical specimens.

Paecilomyces spp incubated at 30°C for 72 hours was the most optimal positive control, with numerous yeast and hyphal forms. The fastest staining process involved 2 minutes of periodic acid and Schiff reagent and 10 dips of light green solution. Tap water was as effective as distilled water. Validation was successfully achieved. Clinical cases all stained identical to formalin-fixed, paraffin-embedded tissue stained with hematoxylin-eosin and periodic acid-Schiff.

Ultra-rapid periodic acid-Schiff provides fast and reliable identification of fungal organisms on fresh tissue. Development of a concurrent positive control allows for quality control and validation.