Cancer Diagnosis Program | Biorepositories and Biospecimen Research Branch

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DNA extraction from FFPE tissue samples - a comparison of three procedures.

Author(s): Sarnecka AK, Nawrat D, Piwowar M, Ligęza J, Swadźba J, Wójcik P

Publication: Contemp Oncol (Pozn), 2019, Vol. 23, Page 52-58

PubMed ID: 31061638 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this study was to investigate the effects of DNA extraction kit choice on the yield and purity of DNA from archival FFPE blocks. Forty-two archival FFPE blocks containing melanoma, colon cancer, or lung adenocarcinoma were included in the study but no processing or storage details were provided. Fresh 5 µm sections were cut from each block and a representative slide was H&E stained. Unstained sections were deparaffinized in xylene and tumor-rich areas were microdissected from the slide using the H&E section as a guide. DNA was extracted from equal amounts of tissue using the QIAamp DNA FFPE Tissue Kit, the Cobas DNA Sample Preparation Kit, or the Maxwell Kit according to the manufacturer’s directions. DNA was quantified spectrophotometrically using a NanoDrop and using the double-stranded Qubit 3.0 Kit. DNA purity was evaluated based on spectrophotometric ratios.

   Summary of Findings:

   The average DNA concentration was significantly lower when extraction was with the QIAamp Kit than the Cobas Kit (using spectrophotometer 18.00 versus 50.50 ng/µL, P<0.0001 and using Qubit 4.79 versus 9.15 ng/µL, P=0.0009) or MAXwell Kit (using spectrophotometer 18.00 versus 102.72 ng/µL, P<0.0001 and using Qubit 4.79 versus 31.28 ng/µL, P=0.0120), but levels were modestly correlated between the QIAamp and other extraction methods (ρ=0.51-0.71). While concentrations were generally higher when extraction was with the Maxwell Kit than the Cobas Kit, the differences were not significant and levels were strongly correlated between methods when quantified by spectrophotometer (ρ=0.85) or Qubit (ρ=0.77). The mean ratio of absorbance at 260 to 280 was lower when extracted with the QIAamp Kit than the Cobas Kit (1.78 versus 1.84, P=0.0003) or the Maxwell Kit (1.78 versus 1.82, P=0.0033), but the median ratio was highest when extraction was with the QIAamp Kit (1.90 versus 1.77 with Cobas and 1.81 with Maxwell).

   Biospecimens

   • Tissue - Colorectal
   • Tissue - Skin
   • Tissue - Lung

   Preservative Types

   • Formalin

   Diagnoses:

   • Neoplastic - Carcinoma
   • Neoplastic - Melanoma

   Platform:

   Analyte	Technology Platform
   DNA	Spectrophotometry
   DNA	Fluorometry

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Spectrophotometry Specific	Technology platform	NanoDrop Qubit 3.0
   Analyte Extraction and Purification	Analyte isolation method	Cobas DNA Sample Preparation Kit QIAamp DNA FFPE Tissue Kit Maxwell 16 FFPE Plus LEV DNA Purification Kit

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