Laboratory investigation; a journal of technical methods and pathology · 2026
Optimizing DNA Extraction from Archival Prostate Biopsy Samples.
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Amaral Adrianna, DePaula Oliveira Lia, Dairo Oluwademilade, Villanueva-Pachas Clarence Rachel, Feng Kewen, Woo Juhyung, Lu Jiayun, Beebe-Dimmer Jennifer L, Thrift Aaron P, Wu Xiao-Cheng, Stroup Antoinette, Shrestha Anshu, Gomez Scarlett Lin, Pollock Sam, Yacoub Rami, Ward Kevin C, Joshu Corinne, Huang Franklin, Hamilton Ann S, Haiman Christopher A, Lotan Tamara L
Аннотация
PURPOSE: In this paper, we compare the performance of three distinct nucleic acid extraction protocols in a large population-based prostate cancer cohort, aiming to optimize DNA yield from limited tumor samples in prostate needle biopsies.
MATERIAL AND METHODS: We utilized two commercially available extraction kits and identified the minimum digitally measured tumor area yielding sufficient DNA for sequencing panels (defined as >50 ng of total DNA). Representative FFPE prostate cancer biopsies were processed from 643 patients recruited through population-based U.S. cancer registries as part of the multi-institutional RESPOND study. Scanned whole slide images were used to annotate and measure the tumor region prior to manual macrodissection and DNA isolation using the extraction kits.
RESULTS: The QIAamp® FFPE Advanced® kit resulted in the highest median DNA yield compared to combined RNA/DNA or DNA extraction using the AllPrep® kit (4.3 vs 2.1 or 2.1 ng per mm2 total tumor, p=0.0004 and p<0.0001, respectively). DNA yield significantly correlated with total tumor area for both kits. Using a tumor area cutoff of 2 mm2 for the Advanced® kit, or 5 mm2 for the AllPrep® DNA/RNA kit, with macrodissection across ten 4 μm tissue sections, 86% to 94% of samples generated more than 50 ng of total DNA. Both kits resulted in comparable panel-based sequencing quality metrics, including percentage duplicated reads and median insert length. For DNA quantity and median target coverage at sequencing, the Advanced® kit outperformed the AllPrep® kit, but it does not allow simultaneous RNA extraction.
CONCLUSIONS: For a DNA yield of at least 50 ng, we suggest a cutoff of 2 mm2 of tumor area with ten 4 μm macrodissected tissue sections for a prostate cancer needle core biopsy to be considered sufficient for DNA extraction.
MATERIAL AND METHODS: We utilized two commercially available extraction kits and identified the minimum digitally measured tumor area yielding sufficient DNA for sequencing panels (defined as >50 ng of total DNA). Representative FFPE prostate cancer biopsies were processed from 643 patients recruited through population-based U.S. cancer registries as part of the multi-institutional RESPOND study. Scanned whole slide images were used to annotate and measure the tumor region prior to manual macrodissection and DNA isolation using the extraction kits.
RESULTS: The QIAamp® FFPE Advanced® kit resulted in the highest median DNA yield compared to combined RNA/DNA or DNA extraction using the AllPrep® kit (4.3 vs 2.1 or 2.1 ng per mm2 total tumor, p=0.0004 and p<0.0001, respectively). DNA yield significantly correlated with total tumor area for both kits. Using a tumor area cutoff of 2 mm2 for the Advanced® kit, or 5 mm2 for the AllPrep® DNA/RNA kit, with macrodissection across ten 4 μm tissue sections, 86% to 94% of samples generated more than 50 ng of total DNA. Both kits resulted in comparable panel-based sequencing quality metrics, including percentage duplicated reads and median insert length. For DNA quantity and median target coverage at sequencing, the Advanced® kit outperformed the AllPrep® kit, but it does not allow simultaneous RNA extraction.
CONCLUSIONS: For a DNA yield of at least 50 ng, we suggest a cutoff of 2 mm2 of tumor area with ten 4 μm macrodissected tissue sections for a prostate cancer needle core biopsy to be considered sufficient for DNA extraction.