The Critical Role of Preanalytical Factors in Validating Cytology Specimens for Hybrid-Capture…

Cohort studyThe Journal of molecular diagnostics : JMD· Jul 2026

The Critical Role of Preanalytical Factors in Validating Cytology Specimens for Hybrid-Capture Next-Generation Sequencing: From Smear to Sequence.

Holly Atwell, Sinchita Roy-Chowdhuri, Rana Seyedjafari … Gloria Hopkins Sura

CytopathologyCohort studyQ1IF 3.5

Papanicolaou-stained smears show markedly lower next-generation sequencing target coverage than FFPE tissue

The Critical Role of Preanalytical Factors in Validating Cytology Specimens for Hybrid-Capture Next-Generation Sequencing: From Smear to Sequence.

Holly Atwell … Gloria Hopkins Sura

The Journal of molecular diagnostics · Jul 2026 · Q1 in Pathology

Background

Cytologic specimens, often the only available sample, are increasingly relied upon for comprehensive genomic profiling in patients with advanced malignancies, necessitating detailed characterization of next-generation sequencing (next-generation sequencing) assay performance.

Methods

This study retrospectively analyzed 10,900 unique specimens sequenced on a custom next-generation sequencing panel, including a focused subset of 455 consecutive cytologic specimens and 2759 FFPE surgical pathology controls.

n = 10,900 unique specimens

Results

Only 74.7% of target regions hit adequate sequencing depth in these smears, versus ~98% for FFPE.

98%

Cell blocks/FFPE

74.7%

Papanicolaou-stained s

89.2%

Combined preparations

More results

Specimen types included Diff-Quik-stained smears, Papanicolaou-stained smears, cell-block sections, and FFPE tissue.

Cell blocks performed comparably to FFPE tissue across all quality metrics (median percentage target coverage ≥98%).

“

Conclusion

Although cytologic smears yield DNA suitable for hybrid-capture next-generation sequencing, specimen-type-specific validation criteria and cytology-aware normalization strategies are essential for maximizing clinical yield and reliability.

The Critical Role of Preanalytical Factors in Validating Cytology Specimens for Hybrid-Capture Next-Generation Sequencing: From Smear to Sequence.

The Critical Role of Preanalytical Factors in Validating Cytology Specimens for Hybrid-Capture Next-Generation Sequencing: From Smear to Sequence.

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