scholarly journals Gene-expression analysis of a colorectal cancer-specific discriminatory transcript set on formalin-fixed, paraffin-embedded (FFPE) tissue samples

2015 ◽  
Vol 10 (1) ◽  
Author(s):  
Alexandra Kalmár ◽  
Barnabás Wichmann ◽  
Orsolya Galamb ◽  
Sándor Spisák ◽  
Kinga Tóth ◽  
...  
Keyword(s):  
Gene Expression ◽  
Colorectal Cancer ◽  
Expression Analysis ◽  
Gene Expression Analysis ◽  
Ffpe Tissue ◽  
Tissue Samples ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Formalin Fixed

scholarly journals Maximizing RNA Yield from Archival Renal Tumors and Optimizing Gene Expression Analysis

2009 ◽  
Vol 15 (1) ◽  
pp. 80-85 ◽  
Author(s):  
Sean T. Glenn ◽  
Karen L. Head ◽  
Bin T. Teh ◽  
Kenneth W. Gross ◽  
Hyung L. Kim
Keyword(s):  
Gene Expression ◽  
Expression Analysis ◽  
Reverse Transcription ◽  
Gene Expression Analysis ◽  
Rna Isolation ◽  
Proteinase K ◽  
Specific Primers ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Formalin Fixed

Formalin-fixed, paraffin-embedded tissues are widely available for gene expression analysis using TaqMan ® PCR. Five methods, including 4 commercial kits, for recovering RNA from paraffin-embedded renal tumor tissue were compared. The MasterPure™ kit from Epicentre produced the highest RNA yield. However, the difference in RNA yield between the kit from Epicenter and Invitrogen’s TRIzol method was not significant. Using the top 3 RNA isolation methods, the manufacturers’ protocols were modified to include an overnight Proteinase K digestion. Overnight protein digestion resulted in a significant increase in RNA yield. To optimize the reverse transcription reaction, conventional reverse transcription with random oligonucleotide primers was compared to reverse transcription using primers specific for genes of interest. Reverse transcription using gene-specific primers significantly increased the quantity of cDNA detectable by TaqMan ® PCR. Therefore, expression profiling of formalin-fixed, paraffin-embedded tissue using TaqMan® qPCR can be optimized by using the MasterPure™ RNA isolation kit modified to include an overnight Proteinase K digestion and gene-specific primers during the reverse transcription.

Clinical relevance of Ki67 gene expression analysis using formalin-fixed paraffin-embedded breast cancer specimens

Breast Cancer ◽  
2012 ◽  
Vol 20 (3) ◽  
pp. 262-270 ◽  
Author(s):  
Satoko Yamamoto ◽  
Mutsuko Ibusuki ◽  
Yutaka Yamamoto ◽  
Peifen Fu ◽  
Saori Fujiwara ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Gene Expression ◽  
Expression Analysis ◽  
Clinical Relevance ◽  
Gene Expression Analysis ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Formalin Fixed

scholarly journals A High-Sensitivity, Medium-Density, and Target Amplification–Free Planar Waveguide Microarray System for Gene Expression Analysis of Formalin-Fixed and Paraffin-Embedded Tissue

2009 ◽  
Vol 55 (11) ◽  
pp. 1995-2003 ◽  
Author(s):  
Stephan Schwers ◽  
Elke Reifenberger ◽  
Mathias Gehrmann ◽  
Alexandre Izmailov ◽  
Kerstin Bohmann
Keyword(s):  
Gene Expression ◽  
Correlation Coefficient ◽  
Expression Analysis ◽  
Gene Expression Analysis ◽  
Dynamic Range ◽  
Ffpe Tissue ◽  
Planar Waveguides ◽  
Assay Sensitivity ◽  
Tissue Samples ◽  
Formalin Fixed

Abstract Background: Many microarray platforms and their associated assay chemistries do not work properly with RNA extracted from formalin-fixed, paraffin-embedded (FFPE) tissue samples, a feature that severely hampers the use of microarrays in oncology applications, for which FFPE tissue is the routine specimen. Furthermore, the limited sensitivity of most microarray platforms requires time-consuming and costly amplification reactions of the target RNA, which negatively affects clinical laboratory work flow. Methods: We developed an approach for sensitively and reliably measuring mRNA abundances in FFPE tissue samples. This approach involves automated RNA extractions, direct hybridization of extracted RNA to immobilized capture probes, antibody-mediated labeling, and readout with an instrument applying the principle of planar waveguides (PWG). A 14-gene multiplex assay conducted with RNA isolated from 20 FFPE blocks was correlated to an analysis of the same with reverse-transcription quantitative real-time PCR (RT-qPCR). Results: The assay sensitivity for gene expression analysis obtained for the PWG microarray platform was <10 fmol/L, eliminating the need for target preamplification. We observed a correlation coefficient of 0.87 to state-of-the-art RT-qPCR technology with RNA isolated from FFPE tissue, despite a compressed dynamic range for the PWG system (a 2.9-log dynamic range for PWG in our test system vs 5.0 logs for RT-qPCR). The precision of the PWG platform was comparable to RT-qPCR (Pearson correlation coefficient of 0.9851 for PWG vs 0.9896 for RT-qPCR) for technical replicates. Conclusions: The presented PWG platform demonstrated excellent sensitivity and precision and is especially well suited for any application for which fast, simple, and robust multiplex assays of RNA in FFPE tissue are required.

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