Optimizing fixation and staining to reduce cell detachment in wet-fixed direct smears, a quantitative evidence-based quality improvement study
Abstract
Wet-fixed cytologic smears are inherently prone to cell detachment, which causes cross-contamination and reduces smear cellularity, potentially leading to false-positive diagnoses and diagnostic uncertainty. This study evaluated the effects of the intermediate alcohol step, fixation duration, and slide type on detachment to establish an evidence-based optimized protocol. We evaluated: (i) the necessity and optimal concentration of the intermediate alcohol step; (ii) fixation duration from 30 min to 24 h; and (iii) slide type (ordinary vs. positively charged). Detachment was scored semi-quantitatively using a streak-wiping method. Wilcoxon and Friedman tests were used for paired and block comparisons, respectively. The intermediate alcohol step significantly reduced detachment ( p < 0.05), with 50% ethanol superior to 75% ( p = 0.012). Prolonged fixation progressively decreased detachment ( p < 0.001), with no further statistically significant reduction beyond 3 h. Under optimized conditions, ordinary and positively charged slides performed comparably ( p = 0.811). An evidence-based strategy for laboratory quality improvement—extending fixation toward 3 h, including an intermediate alcohol step (preferably 50% ethanol) at each transition between water and 95% ethanol, and using ordinary slides—substantially reduces detachment. By preserving cellularity and helping to reduce the risk of cross-contamination, this protocol offers an implementable quality improvement tool for cytology laboratories, potentially enhancing diagnostic reliability and patient safety in wet-fixed cytologic preparations.
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BibTeX
@article{zou2026optimizing,
title = {{Optimizing fixation and staining to reduce cell detachment in wet-fixed direct smears, a quantitative evidence-based quality improvement study}},
author = {Yinying Zou and Xin Wang and Yan Chen and Zihan Wu},
journal = {Scientific Reports},
year = {2026},
month = oct,
doi = {10.1038/s41598-026-76374-3},
url = {https://www.nature.com/articles/s41598-026-76374-3},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Optimizing fixation and staining to reduce cell detachment in wet-fixed direct smears, a quantitative evidence-based quality improvement study
AU - Yinying Zou
AU - Xin Wang
AU - Yan Chen
AU - Zihan Wu
PY - 2026
DA - 2026/10/17
JO - Scientific Reports
DO - 10.1038/s41598-026-76374-3
UR - https://www.nature.com/articles/s41598-026-76374-3
AB - Wet-fixed cytologic smears are inherently prone to cell detachment, which causes cross-contamination and reduces smear cellularity, potentially leading to false-positive diagnoses and diagnostic uncertainty. This study evaluated the effects of the intermediate alcohol step, fixation duration, and slide type on detachment to establish an evidence-based optimized protocol. We evaluated: (i) the necessity and optimal concentration of the intermediate alcohol step; (ii) fixation duration from 30 min to 24 h; and (iii) slide type (ordinary vs. positively charged). Detachment was scored semi-quantitatively using a streak-wiping method. Wilcoxon and Friedman tests were used for paired and block comparisons, respectively. The intermediate alcohol step significantly reduced detachment ( p < 0.05), with 50% ethanol superior to 75% ( p = 0.012). Prolonged fixation progressively decreased detachment ( p < 0.001), with no further statistically significant reduction beyond 3 h. Under optimized conditions, ordinary and positively charged slides performed comparably ( p = 0.811). An evidence-based strategy for laboratory quality improvement—extending fixation toward 3 h, including an intermediate alcohol step (preferably 50% ethanol) at each transition between water and 95% ethanol, and using ordinary slides—substantially reduces detachment. By preserving cellularity and helping to reduce the risk of cross-contamination, this protocol offers an implementable quality improvement tool for cytology laboratories, potentially enhancing diagnostic reliability and patient safety in wet-fixed cytologic preparations.
SN - 2045-2322
ER -