Loss of heterozygosity studies revisited: prior quantification of the amplifiable DNA content of archival samples improves efficiency and reliability.

Author(s): Farrand K, Jovanovic L, Delahunt B, McIver B, Hay ID, Eberhardt NL, Grebe SK

Publication: J Mol Diagn, 2002, Vol. 4, Page 150-8

PubMed ID: 12169676 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this study was to determine the effects of formalin fixation and snap-freezing on the ratio of amplifiable DNA found in thyroid tumor tissue.

   Summary of Findings:

   Raw DNA concentrations extracted from FFPE specimens were significantly lower than those extracted from frozen tumor specimens and their matched buffy coat specimens. Most of the raw input DNA was amplifiable from frozen specimens with ratios of amplifiable DNA to input DNA ranging between 1:1 and 1:5. However, a much smaller proportion of input DNA from FFPE specimens was amplifiable with ratios ranging from 1:5 to 1:3625. The longer amplicon failed to amplify in 14 out of 20 FFPE-derived specimens while it successfully amplified in all 10 frozen tissue-derived specimens. Use of the assay for the ABR locus, which was the smallest amplicon, led to higher ratio of amplifiable DNA to input DNA.

   Biospecimens

   • Tissue - Thyroid Gland
   • Bodily Fluid - Buffy Coat

   Preservative Types

   • Formalin
   • Frozen

   Diagnoses:

   • Neoplastic
   • Neoplastic - Normal Adjacent

   Platform:

   Analyte	Technology Platform
   DNA	Real-time qPCR
   DNA	Fluorometry

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Biospecimen Preservation	Type of fixation/preservation	Formalin (buffered) Snap frozen
   Biospecimen Aliquots and Components	Blood and blood products	Buffy coat
   Biospecimen Acquisition	Biospecimen location	Blood Thyroid
   Real-time qPCR Specific	Length of gene fragment	80-120 bp 175-215 bp 280-322 bp

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