ArticleScientific reports2016
Straightforward and sensitive RT-qPCR based gene expression analysis of FFPE samples.
Article in Scientific reports, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 23 papers.
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Who cites it
23 citing papers in PubMed.
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- Cutavirus Infection in Large-Plaque Parapsoriasis, a Premalignant Condition of Mycosis Fungoides.The Journal of infectious diseases · 2024Article
- Streamlined Full-Length Total RNA Sequencing of Paraformaldehyde-Fixed Brain Tissues.International journal of molecular sciences · 2024Article
- Analytical validation and algorithm improvement of HepatoPredict kit to assess hepatocellular carcinoma prognosis before a liver transplantation.Practical laboratory medicine · 2024Article
- mRNA Capture Sequencing and RT-qPCR for the Detection of Pathognomonic, Novel, and Secondary Fusion Transcripts in FFPE Tissue: A Sarcoma Showcase.International journal of molecular sciences · 2022Article
- Comprehensive evaluation of the test for 5'-/3'-end mRNA unbalanced expression as a screening tool for ALK and ROS1 fusions in lung cancer.Cancer medicine · 2022Article
- Expression of CD274 mRNA Measured by qRT-PCR Correlates With PD-L1 Immunohistochemistry in Gastric and Urothelial Carcinoma.Frontiers in oncology · 2022Article
- A comprehensive tool box for large animal studies of intervertebral disc degeneration.JOR spine · 2021Article
- Sensitive detection of colorectal cancer in peripheral blood by a novel methylation assay.Clinical epigenetics · 2021Article
- Effects of Essential Oils-Based Supplement andAnimals : an open access journal from MDPI · 2021Article
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- Germline mutations in the new E1' cryptic exon of theJournal of medical genetics · 2020Article
- Selection of endogenous control genes for normalising gene expression data derived from formalin-fixed paraffin-embedded tumour tissue.Scientific reports · 2020Article
- Comprehensive analysis of soluble RNAs in human embryo culture media and blastocoel fluid.Journal of assisted reproduction and genetics · 2020Article
- BDNF Expression in Cortical GABAergic Interneurons.International journal of molecular sciences · 2020Article
- Examination of the Expression of Immunity Genes and Bacterial Profiles in the Caecum of Growing Chickens Infected withAnimals : an open access journal from MDPI · 2019Article
- Systematic evaluation of RNA quality, microarray data reliability and pathway analysis in fresh, fresh frozen and formalin-fixed paraffin-embedded tissue samples.Scientific reports · 2018Article
- MicroRNA-196a-5p is a potential prognostic marker of delayed lymph node metastasis in early-stage tongue squamous cell carcinoma.Oncology letters · 2018Article
- Multiplexed analysis of fixed tissue RNA using Ligation in situ Hybridization.Nucleic acids research · 2017Article
Corrections and comments
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Authors and funding
6 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Fragmented RNA from formalin-fixed paraffin-embedded (FFPE) tissue is a known obstacle to gene expression analysis. In this study, the impact of RNA integrity, gene-specific reverse transcription and targeted cDNA preamplification was quantified in terms of reverse transcription polymerase chain reaction (RT-qPCR) sensitivity by measuring 48 protein coding genes on eight duplicate cultured cancer cell pellet FFPE samples and twenty cancer tissue FFPE samples. More intact RNA modestly increased gene detection sensitivity by 1.6 fold (earlier detection by 0.7 PCR cycles, 95% CI = 0.593-0.850). Application of gene-specific priming instead of whole transcriptome priming during reverse transcription further improved RT-qPCR sensitivity by a considerable 4.0 fold increase (earlier detection by 2.0 PCR cycles, 95% CI = 1.73-2.32). Targeted cDNA preamplification resulted in the strongest increase of RT-qPCR sensitivity and enabled earlier detection by an average of 172.4 fold (7.43 PCR cycles, 95% CI = 6.83-7.05). We conclude that gene-specific reverse transcription and targeted cDNA preamplification are adequate methods for accurate and sensitive RT-qPCR based gene expression analysis of FFPE material. The presented methods do not involve expensive or complex procedures and can be easily implemented in any routine RT-qPCR practice.
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