Gene expression profiling of FFPE breast cancer samples
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Maurizio Callari
PROVIDER: E-GEOD-38554 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:The use of Affymetrix U133 2.0 Plus chips on FFPE samples when coupled with a qPCR-based sample pre-assessment step, yielded satisfactory results from the point of view of biological reliability. When compared with the Illumina DASL WG platform, specifically designed for degraded RNA, the data generated with the Affymetrix platform showed a wider interquartile range (IQR 1.32 vs 0.57, p<2.2x10-16) suggesting a superior discriminatory power within samples as indicated by the good agreement with the immunohistiochemically derived ER status. FFPE primary breast cancer samples profiled using Affymetrix HG-U133 Plus 2.0 microarray platform after RNA amplification with the Nugen WT-Ovation FFPE System The following criteria were considered for a direct comparison of 12 GEPs obtained from Affymetrix and DASL platforms: gene variability as defined by IQR, ESR1 expression in ER status subgroups defined by IHC, distribution of fold changes for predefined ER related genes when comparing ER positive and negative samples.
2015-02-25 | E-GEOD-51450 | biostudies-arrayexpress
Project description:This study was performed to evaluated RNA extraction and gene expression analysis of FFPE specimen stored for more than 20 years. Using long time stored FFPE material; large retrospective studies correlating molecular features with therapeutic response and clinical outcome, can be performed. Quantitative PCR (qPCR) was used to evaluate RNA extraction methods and to compare gene expression profiles of FFPE and fresh frozen (FF) tissue. Extracted RNA was subsequently subjected to microarray analysis and compared to qPCR data. The Ambion RecoverAll kit appears to be particularly suited for RNA extraction of long time stored FFPE tissues. Gene expression analysis using Affymetrix platform displayed a high degree of correlation for endogenous control genes comparing FF and FFPE tissues. We conclude that high quality gene expression signatures can be recognized using Affymetrix gene expression platform on FFPE tissue stored for more than 20 years. However, a general interpretation must be done with caution as different FFPE procedures have varying effects on RNA quality. Three RNA extraction methods from Roche (Basel, Switzerland), Ambion (Austin, TX, USA) and Qiagen (Hilden, Germany), designed for FFPE material was used. The methods were compared using qPCR. For the qPCR analysis, two different concentrations of input cDNA were used (20ng and 100ng) and 32 human endogenous control genes were examined. RNA from the Ambion FFPE kit was further analyzed by using microarray. Amplification of RNA prior to microarray analysis was performed using Nugen technologies (San Carlos, CA, USA). Nugen has developed amplification kits both for FFPE and FF materials: WT- ovation FFPE RNA amplification System V2 (Nugen FFPE), Ovation FF RNA Amplification System V2 (Nugen V2 FF) and Ovation FF Pico RNA Amplification System (Nugen PICO FF). The Nugen FFPE kit, designed for FFPE material was only used for FFPE tissue. The Nugen Pico FF kit, designed to target small amounts of FF RNA (>500 pg) and the Nugen V2 FF kit designed to target total FF RNA, were only used for FF tissue. Affymetrix standard amplification protocol (Affy FF) designed for FF RNA was also included as the standard method for amplification.
2011-09-14 | E-GEOD-31735 | biostudies-arrayexpress
Project description:The use of Affymetrix U133 2.0 Plus chips on FFPE samples when coupled with a qPCR-based sample pre-assessment step, yielded satisfactory results from the point of view of biological reliability. When compared with the Illumina DASL WG platform, specifically designed for degraded RNA, the data generated with the Affymetrix platform showed a wider interquartile range (1.32 vs 0.57, p<2.2x10-16) suggesting a superior discriminatory power within samples as indicated by the good agreement with the immunohistiochemically derived ER status. FFPE primary breast cancer samples profiled using Illumina DASL WG platform after RNA amplification with the Nugen WT-Ovation FFPE System
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Project description:The use of Affymetrix U133 2.0 Plus chips on FFPE samples when coupled with a qPCR-based sample pre-assessment step, yielded satisfactory results from the point of view of biological reliability. When compared with the Illumina DASL WG platform, specifically designed for degraded RNA, the data generated with the Affymetrix platform showed a wider interquartile range (1.32 vs 0.57, p<2.2x10-16) suggesting a superior discriminatory power within samples as indicated by the good agreement with the immunohistiochemically derived ER status. FFPE primary breast cancer samples profiled using Illumina DASL WG platform after RNA amplification with the Nugen WT-Ovation FFPE System The following criteria were considered for a direct comparison of 12 GEPs obtained from Affymetrix and DASL platforms: gene variability as defined by IQR, ESR1 expression in ER status subgroups defined by IHC, distribution of fold changes for predefined ER related genes when comparing ER positive and negative samples.
2015-02-25 | E-GEOD-51451 | biostudies-arrayexpress
Project description:To study feasibility of gene expression profiling from FFPE tissues using NuGen amplified mRNA hybridized on Affymetrix GeneChip Human Gene 1.0 ST arrays, we designed a pilot study utilizing samples from prostate cancer cohort. We selected samples from large-scale epidemiologic studies and clinical trials representative of a wide variety of fixation times, block ages and block storage conditions.
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Project description:To study feasibility of gene expression profiling from FFPE tissues using NuGen amplified mRNA hybridized on Affymetrix GeneChip Human Gene 1.0 ST arrays, we designed a pilot study utilizing samples from prostate cancer cohort. We selected samples from large-scale epidemiologic studies and clinical trials representative of a wide variety of fixation times, block ages and block storage conditions.
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| EGAD00001008699 | EGA