Revisiting an old technique: rehydration of air-dried slides is a viable technique for Pap-stained fine needle aspiration slides for novel situations

Introduction: Traditional Pap staining procedure involves immediate ethanol-fixation of prepared smears. When our department absorbed cytopathology services of an affiliate hospital using nonspecialized assistants, we had problems with poorly fixed slides. We evaluated an alternative approach of rehydration of air-dried slides in comparison to both immediate and delayed alcohol fixation for Pap staining. Materials and methods: Fine needle aspirations were performed on deidentified unfixed autopsy tissue. Sampled tissues included lung, liver, thyroid, breast, and a mediastinal mass. Paired aspirate smears were immediately fixed, air-dried and rehydrated before staining, or delayed fixation after a 5-minute air dry (DF). Additional smears were Diff-Quik stained for comparison. Slides were Pap-stained at 0 days, 1 day, 3 days, or 5 days postaspiration and numerically scored for cytomorphologic feature quality. Data were compared with one-way analysis of variance analysis. Results: We evaluated 237 pairs of smears. The immediately fixed and air-dried groups displayed higher quality for nuclear borders, nuclear detail, distinct cell borders, and cytoplasmic staining at every time point (all P values <0.05) compared to the DF group. Rehydration did not reverse the air-dry artifact from delayed fixation in the DF group. Conclusions: Air-drying with delayed rehydration/alcohol fixation maintains diagnostic quality in settings for which traditional preparative techniques are not optimal, technical assistance is limited, and transport is problematic. Our series demonstrates that air-dried slides maintained high quality when processed up to 5 days postaspiration. We demonstrate a technique that improves pathology outreach by providing better quality diagnostic material with minimal additional personnel costs.