NGS from FFPE Samples

Otogenetics developed QC standards and classification of sample grades for FFPE exome sequencing

QC standards and classification of FFPE gDNA samples based on initial sample quality.

Otogenetics uses gel electrophoresis and spectrophotometer assays to assess the quality of initial gDNA. DNA from fresh tissues appears as a large molecular weight band above 10Kb on the gel. gDNA from FFPE samples have various appearances on the gel depending on how fixation was done, how gDNA extraction was performed, and how well gDNAs were stored after purification. The best ones appear as smears across from low to high molecular weight species, some appear as medium-size smears, and some as low molecular weight smears only. QC of gDNA from FFPE tissues were categorized into Grades 1, 2, and 3 based on original smear appearance and PCR QC, as demonstrated in our downloadable white paper.

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Otogenetics adjusts workflow to achieve maximized completion of FFPE exome sequencing.

Among FFPE gDNA samples submitted for whole exome sequencing, mostly were classified as Grade 2. For Grade 1 FFPE gDNAs, all of the samples achieved satisfactory sequencing results. For Grade 2 FFPE gDNAs, 83% of the samples are expected to achieve satisfactory sequencing. For Grade 3 gDNAs, 75% are expected to achieve satisfactory sequencing. Adjustment of the workflow ensures the achievement of desired coverage; however, the duplication rate is higher in lower grade FFPE gDNA. Sequencing data quality itself (Q30) is not affected by the quality of gDNA from FFPE tissues.

Please download our white paper to learn more or contact Otogenetics for a customized workflow dedicated specifically for your FFPE samples.