Fixation Effects on Variant Calling in a Clinical Resequencing Panel

Area of Science:

• Pathology
• Molecular Biology
• Genomics

Background:

• Formalin fixation is standard for tissue preservation but introduces artifacts impacting molecular assay accuracy.
• These artifacts can compromise the reliability of diagnostic and genetic testing results.

Purpose of the Study:

• To evaluate alternative tissue fixatives for improved performance in next-generation sequencing (NGS) assays.
• To assess if alternative fixatives maintain adequate tissue morphology for pathological diagnosis.
• To compare the impact of different fixatives on sequencing quality and variant calling.

Main Methods:

• Prospective study utilizing a clinical-grade, targeted resequencing assay.
• Analysis of tumor and normal colon epithelial tissues preserved with formalin and an alternative fixative.
• Quantification of sequencing quality metrics and variant calling accuracy.

Main Results:

• An alternative fixative was identified that significantly reduces formalin-induced artifacts.
• The alternative fixative demonstrated comparable or improved performance in NGS assays.
• Adequate tissue morphology for pathological review was retained using the alternative fixative.

Conclusions:

• Alternative fixatives offer a viable solution to mitigate formalin-related artifacts in molecular testing.
• Optimized tissue fixation can enhance the accuracy of next-generation sequencing results.
• This finding supports the adoption of improved fixation methods for integrated diagnostic workflows.