Table 1 Common pre-analytical and sample preparation issues related to different sample types
From: Technological considerations for genome-guided diagnosis and management of cancer
Sample type | Common issues | Impact | Contingencies/solutions |
|---|---|---|---|
Formalin-fixed, paraffin-embedded (FFPE) | • Low yield of DNA • DNA degradation • DNA base modification • RNA degradation | • Reduced complexity libraries; library failure; decreased sensitivity • Reduced complexity; library failure; decreased sensitivity • Increased false positive rate • Library failure; high duplication | • DNA repair; pooling of indexed libraries prior to capture (exomes or panels); specialized low input library methods • DNA repair; short amplicon amplification; specialized library methods • FFPE-aware filtering of variants; DNA repair • Selection-based or targeted preparation instead of polyA-based preparation |
Fresh frozen tissue of bulk cells | • Buffer or process-induced modification of DNA bases | • Increased false positive rate | • Chelation of oxidative species; oxidation aware filtering |
Single cells | • Low DNA yield • Whole genome amplification (WGA) bias • Low RNA yield | • Library failure • Increased false positives and false negatives • Library failure | • WGA • Optimized WGA • Whole transcriptome amplification (WTA) |
Liquid biopsy | • Low DNA yield of cfDNA • Low purity of ctDNA in cfDNA • Low DNA yield from CTCs • Low RNA yield and quality from CTCs • Low RNA yield from EVs | • Library failure; reduced sensitivity • Reduced sensitivity • Library failure; reduced sensitivity; reduced specificity • Library failure • Library failure | • Optimized library preparation; specialized library preparation • High sequencing depth; molecular barcoding (UMIs) • WGA • WTA; specialized library preparation. • WTA; specialized library preparation. |