Suitability of melanoma FFPE samples for NGS libraries: time and quality thresholds for downstream molecular tests. (9th August 2018)

• Record Type: Journal Article
• Authors: Millán-Esteban, David; Reyes-García, Diana; García-Casado, Zaida; Bañuls, José; López-Guerrero, José Antonio; Requena, Celia; Rodríguez-Hernández, Aranzazu; Traves, Víctor; Nagore, Eduardo
• Is Part Of: Biotechniques. Volume 65:Number 2(2018)
• Page Start: 79
• Page End: 85
• Publication Date: 2018-08-09
• Subjects: DIN -- FFPE -- library -- melanoma -- NGS -- QC -- quality -- threshold -- time; Biology, Experimental -- Periodicals; Molecular biology -- Periodicals; Medical technology -- Periodicals; Biology, Experimental; Medical technology; Molecular biology; Clinical Laboratory Techniques -- Periodicals; Research -- Periodicals; Medical Laboratory Science -- Periodicals; Periodicals; Electronic journals; 570
• DOI: 10.2144/btn-2018-0016
• Languages: English
• ISSNs: 0736-6205

Abstract

The use of NGS in clinical practice for precision diagnosis requires a quality starting material. Despite the broadly established use of formalin-fixed paraffin-embedded (FFPE) samples in molecular testing, these usually have low-quality DNA. We established a method to determine the suitability of melanoma FFPE samples for an amplicon-based NGS custom panel analysis. DNA was extracted from unstained melanoma samples and wide local excision samples. Amplicon-based libraries were constructed and tested using time and quality parameters as variables. Time elapsed from sample retrieval >7 years, a quality control value > 5.63 and a DNA integrity value < 2.05 indicated samples were not suitable. A decision tree is provided with rate of samples suitable for analysis according to the combination of these parameters.