Methods comparison for high-resolution transcriptional analysis of archival material on Affymetrix Plus 2.0 and Exon 1.0 microarrays.
Author(s): Linton K, Hey Y, Dibben S, Miller C, Freemont A, Radford J, Pepper S
Publication: Biotechniques, 2009, Vol. 47, Page 587-96
PubMed ID: 19594443 PubMed Review Paper? No
Purpose of Paper
Conclusion of Paper
Studies
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Study Purpose
The purpose of this study was to determine the effects of fixation, RNA extraction kit, labeling method, and array type on microarray analysis of leiomyosarcomas, liposarcomas and synovial sarcomas.
Summary of Findings:
Although RNA of similar quantity and quality was extracted with the Ambion Optimum and QIagen RNEasy kits, the percentage of positive calls on the U133 arrays was higher for samples extracted with the Ambion Optimum kit than samples extracted using the Qiagen RNEasy kit. The percentage of positive calls was also approximately 20% higher for RNA labeled with NuGen FFPE than with Affymetrix One-cycle or Affymetrix Exon WT kits. Interestingly there was no 3' bias in the specimens labeled with the Affymetrix one-cycle kit. When frozen specimens were compared to FFPE specimens, 20% of the positive calls from frozen specimens were not detected in FFPE specimens using the ideal protocol on U133 arrays. When FFPE RNA was hybridized to Exon arrays instead of U133 arrays, sensitivity increased to 92%.
Biospecimens
Preservative Types
- Formalin
- Frozen
Diagnoses:
- Neoplastic - Sarcoma
Platform:
Analyte Technology Platform RNA Automated electrophoresis/Bioanalyzer RNA DNA microarray Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Analyte Extraction and Purification Analyte isolation method Ambion Optimum FFPE
QIAgen RNeasy FFPE
Trizol
DNA microarray Specific Type of array Exon 1.0
U133 Plus 2.0
Biospecimen Preservation Type of fixation/preservation Formalin (buffered)
Frozen
DNA microarray Specific Detection method NuGen
Affymetrix one-cycle
Affymetrix two-cycle
Affymetrix WT Exon