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- Title
Whole-Genome Gene Expression Profiling of Formalin-Fixed, Paraffin-Embedded Tissue Samples.
- Authors
April, Craig; Klotzle, Brandy; Royce, Thomas; Wickham-Garcia, Eliza; Boyaniwsky, Tanya; Izzo, John; Cox, Donald; Jones, Wendell; Rubio, Renee; Holton, Kristina; Matulonis, Ursula; Quackenbush, John; Jian-Bing Fan
- Abstract
Background: We have developed a gene expression assay (Whole-Genome DASLH), capable of generating whole-genome gene expression profiles from degraded samples such as formalin-fixed, paraffin-embedded (FFPE) specimens. Methodology/Principal Findings: We demonstrated a similar level of sensitivity in gene detection between matched freshfrozen (FF) and FFPE samples, with the number and overlap of probes detected in the FFPE samples being approximately 88% and 95% of that in the corresponding FF samples, respectively; 74% of the differentially expressed probes overlapped between the FF and FFPE pairs. The WG-DASL assay is also able to detect 1.3-1.5 and 1.5-2 -fold changes in intact and FFPE samples, respectively. The dynamic range for the assay is ∼3 logs. Comparing the WG-DASL assay with an in vitro transcription-based labeling method yielded fold-change correlations of R² ∼0.83, while fold-change comparisons with quantitative RT-PCR assays yielded R²∼0.86 and R²∼0.55 for intact and FFPE samples, respectively. Additionally, the WGDASL assay yielded high self-correlations (R²>0.98) with low intact RNA inputs ranging from 1 ng to 100 ng; reproducible expression profiles were also obtained with 250 pg total RNA (R²,&#x223C0.92), with ∼71% of the probes detected in 100 ng total RNA also detected at the 250 pg level. When FFPE samples were assayed, 1 ng total RNA yielded self-correlations of R²∼0.80, while still maintaining a correlation of R²∼0.75 with standard FFPE inputs (200 ng). Conclusions/Significance: Taken together, these results show that WG-DASL assay provides a reliable platform for genome-wide expression profiling in archived materials. It also possesses utility within clinical settings where only limited quantities of samples may be available (e.g. microdissected material) or when minimally invasive procedures are performed (e.g. biopsied specimens).
- Subjects
TISSUE analysis; GENE expression; GENETIC regulation; HUMAN genome; GENOMICS; GENETIC research
- Publication
PLoS ONE, 2009, Vol 4, Issue 12, p1
- ISSN
1932-6203
- Publication type
Article
- DOI
10.1371/journal.pone.0008162