Quantitative expression profiling in formalin-fixed paraffin-embedded samples by affymetrix microarrays.

Author(s): Abdueva D, Wing M, Schaub B, Triche T, Davicioni E

Publication: J Mol Diagn, 2010, Vol. 12, Page 409-17

PubMed ID: 20522636 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this study was to compare RNA quality and suitability for microarray analysis of FFPE and frozen kidney, colon and lung tissues and to determine the effects of storage. Different RNA extraction protocols were used for frozen and FFPE specimens.

   Summary of Findings:

   The yield of RNA was significantly lower from FFPE tissue than from frozen specimens (p<0.02). A non-significant decrease in RNA yield was observed with storage both in frozen (2 fold) and FFPE tissue (4 fold). RNA from FFPE tissue was highly degraded (<500 bp) when compared to RNA from frozen specimens. Despite five times more starting RNA, less amplification product was produced from FFPE than frozen tissues (p=0.0004). The amplification yield from FFPE tissue was also 2 fold greater in tissues that had been stored for <1.5 years compared to tissues stored for 7.5 years (p=0.009). When RNA came from frozen tissues, there were 1.4 fold more present calls on the microarray, then when the RNA came from FFPE tissues. Further, the variance was higher from FFPE tissues than from frozen tissues and the raw signal intensity had a smaller range. The signal intensity was reduced in probes detecting the 5' end compared to the 3' end of transcripts from FFPE, but not frozen tissues. Although there was >50% concordance of genes differentially expressed (>2 fold) between tumor and normal tissue for FFPE and frozen specimens, the magnitude of the fold change differed significantly. When only genes with changes of greater than 5 fold were examined, the concordance between FFPE and frozen specimens increased to 90%. Further, the use of more stringent p value cut-offs increased the concordance of differentially expressed genes between tumor and normal tissue in FFPE and frozen specimens.

   Biospecimens

   • Tissue - Kidney
   • Tissue - Lung
   • Tissue - Colorectal

   Preservative Types

   • Formalin
   • Frozen

   Diagnoses:

   • Neoplastic
   • Neoplastic - Normal Adjacent

   Platform:

   Analyte	Technology Platform
   RNA	Spectrophotometry
   RNA	Automated electrophoresis/Bioanalyzer
   RNA	DNA microarray

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Biospecimen Preservation	Type of fixation/preservation	Formalin (buffered) Frozen
   Storage	Storage duration	0.5 years 2.5 years 7.5 years

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