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Rnaeasy ffpe kit

Manufactured by Qiagen
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The RNAeasy FFPE kit is a product designed for the extraction and purification of RNA from formalin-fixed, paraffin-embedded (FFPE) tissue samples. It is a tool used in molecular biology and genomics research.

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Lab products found in correlation

Rnaeasy kit, by Qiagen (3 mentions) 2100 bioanalyzer analysis, by Agilent Technologies (2 mentions) Mirneasy mini kit, by Qiagen (2 mentions) Lightcycler 480, by Roche (2 mentions) Quantitect reverse transcription kit, by Qiagen (2 mentions) Sybr green master mix, by Thermo Fisher Scientific (2 mentions) Pkd buffer, by Qiagen (2 mentions) Nanodrop 2000, by Thermo Fisher Scientific (2 mentions) Agilent 2100 bioanalyzer, by Agilent Technologies (2 mentions) 2100 bioanalyzer, by Agilent Technologies (1 mentions) Superscript first strand synthesis system for rt pcr, by Thermo Fisher Scientific (1 mentions) Qiaamp dna ffpe tissue kit, by Qiagen (1 mentions) Taqman probe assay, by Thermo Fisher Scientific (1 mentions) Taqman universal pcr master mix, by Thermo Fisher Scientific (1 mentions) Genechip hta 2, by Thermo Fisher Scientific (1 mentions) Microtome, by Leica camera (1 mentions) Quantstudio 7 system, by Thermo Fisher Scientific (1 mentions) Taqman reverse transcription reagent, by Thermo Fisher Scientific (1 mentions) 384 well pcr plate, by Thermo Fisher Scientific (1 mentions) Lightcycler 480 sybr green 1 master kit, by Roche (1 mentions)

23 protocols using rnaeasy ffpe kit

1

RNA Extraction from FFPE Samples

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RNA was extracted from FFPE sections of primary tumors and reactive lymph nodes using the FFPE RNA Easy Kit (Qiagen, CA), and from cell lines using the RNA Easy Kit (Qiagen, CA), according to the manufacturer’s instructions. The amount and quality of RNA were evaluated by measuring the OD at 260 nm and the 260/230 and 260/280 ratios using a Nanodrop spectrophotometer (Celbio, Milan, Italy). The quality of RNA was also checked using a Bioanalyzer 2100 (Agilent, CA, USA).
Mundo L., Ambrosio M.R., Raimondi F., Del Porro L., Guazzo R., Mancini V., Granai M., Jim Rocca B., Lopez C., Bens S., Onyango N., Nyagol J., Abinya N., Navari M., Ndede I., Patel K., Paolo Piccaluga P., Bob R., de Santi M.M., Russell R.B., Lazzi S., Siebert R., Stein H, & Leoncini L. (2019). Molecular switch from MYC to MYCN expression in MYC protein negative Burkitt lymphoma cases. Blood Cancer Journal, 9(12), 91.
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2

Comprehensive Characterization of Lung Cancers

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We collected tumor samples from 67 LUAD, 36 LUSC, 9 LCLC patients along with 6 matched normal lung tissues samples following surgery at the University of Michigan. The recruitment of subjects and informed consent were reviewed and approved by our Institutional Review Board. The publically available dataset from TCGA was downloaded using the TCGA portal and the Seoul data from dbGAP. Formalin-fixed, paraffin-embedded (FFPE) sections from 11 adenoid cystic carcinoma samples were from IRCCS AOU San Martino-IST, Genova, Italy. The 24 lung cell lines were purchased from American Type Culture Collection (ATCC) and cultured following their media and growth conditions. Total RNA from frozen tissues or cell lines were isolated using miRNeasy mini kit (Qiagen Valencia, CA) while RNA was isolated from FFPE sections using FFPE RNAeasy kit (Qiagen). Only high quality RNA from frozen sections and cell lines with RNA integrity number (RIN) >8.0, upon 2100 Bioanalyzer analysis (Agilent Santa Clara, CA) were subjected to RNA sequencing (Supplementary Methods).
Dhanasekaran S.M., Balbin O.A., Chen G., Nadal E., Kalyana-Sundaram S., Pan J., Veeneman B., Cao X., Malik R., Vats P., Wang R., Huang S., Zhong J., Jing X., Iyer M., Wu Y.M., Harms P.W., Lin J., Reddy R., Brennan C., Palanisamy N., Chang A.C., Truini A., Truini M., Robinson D.R., Beer D.G, & Chinnaiyan A.M. (2014). Transcriptome Meta-Analysis of Lung Cancer Reveals Recurrent Aberrations in NRG1 and Hippo Pathway Genes. Nature communications, 5, 5893.
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3

Comprehensive Characterization of Lung Cancers

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We collected tumor samples from 67 LUAD, 36 LUSC, 9 LCLC patients along with 6 matched normal lung tissues samples following surgery at the University of Michigan. The recruitment of subjects and informed consent were reviewed and approved by our Institutional Review Board. The publically available dataset from TCGA was downloaded using the TCGA portal and the Seoul data from dbGAP. Formalin-fixed, paraffin-embedded (FFPE) sections from 11 adenoid cystic carcinoma samples were from IRCCS AOU San Martino-IST, Genova, Italy. The 24 lung cell lines were purchased from American Type Culture Collection (ATCC) and cultured following their media and growth conditions. Total RNA from frozen tissues or cell lines were isolated using miRNeasy mini kit (Qiagen Valencia, CA) while RNA was isolated from FFPE sections using FFPE RNAeasy kit (Qiagen). Only high quality RNA from frozen sections and cell lines with RNA integrity number (RIN) >8.0, upon 2100 Bioanalyzer analysis (Agilent Santa Clara, CA) were subjected to RNA sequencing (Supplementary Methods).
Dhanasekaran S.M., Balbin O.A., Chen G., Nadal E., Kalyana-Sundaram S., Pan J., Veeneman B., Cao X., Malik R., Vats P., Wang R., Huang S., Zhong J., Jing X., Iyer M., Wu Y.M., Harms P.W., Lin J., Reddy R., Brennan C., Palanisamy N., Chang A.C., Truini A., Truini M., Robinson D.R., Beer D.G, & Chinnaiyan A.M. (2014). Transcriptome Meta-Analysis of Lung Cancer Reveals Recurrent Aberrations in NRG1 and Hippo Pathway Genes. Nature communications, 5, 5893.
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4

Nucleic Acid Extraction and cDNA Synthesis

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Peripheral-blood DNA/RNA extraction was performed with the MagNA Pure Compact extractor (Roche), according to the manufacturer’s protocol. Tissue DNA/RNA (from normal epithelium, adenoma or tumor sections) was obtained from formalin-fixed paraffin-embedded (FFPE) tissues with a purity greater than 80% as determined by an experienced pathologist. DNA extractions were performed with the QIAamp DNA FFPE Tissue kit (Qiagen N.V.) and RNA extractions with the RNAeasy FFPE kit (Qiagen N.V.). SuperScript First-Strand Synthesis System for RT-PCR (Thermo Fisher Scientific) was used to synthesize cDNA, either from blood or tissue RNA, using random hexamers and oligo-dT, according to the manufacturer’s instructions.
Lorca V., Rueda D., Martín-Morales L., Fernández-Aceñero M.J., Grolleman J., Poves C., Llovet P., Tapial S., García-Barberán V., Sanz J., Pérez-Segura P., de Voer R.M., Díaz-Rubio E., de la Hoya M., Caldés T, & Garre P. (2019). Contribution of New Adenomatous Polyposis Predisposition Genes in an Unexplained Attenuated Spanish Cohort by Multigene Panel Testing. Scientific Reports, 9, 9814.
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5

FFPE RNA Extraction and Cytokine qRT-PCR

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For cytokine mRNA expression studies, RNA was extracted from formalin-fixed paraffin-embedded (FFPE) histological tissues using RNAeasy FFPE kit (Qiagen). Briefly, freshly cut FFPE tissue 5-μm sections were incubated with Qiagen deparaffinization solution prior to RNA purification. Following proteinase K tissue digestion and subsequent incubation at 56°C and 80°C, RNAeasy MinElute spin columns were used to elute RNA, according to the manufacturer’s instructions. Quantitative reverse transcriptase PCR was performed on C1000 Touch Thermal Cycler apparatus (Bio-Rad) using TaqMan universal PCR Master Mix and Taqman probe assays (ThermoFisher Scientific): RPLP0 (Hs99999902_m1); GzmB (Hs01554355_m1); IFNG (Hs99999041_m1); IL10 (Hs00961622_m1); IL12A (Hs009673447_m1); IL21 (Hs00222327_m1). Results were expressed as relative expression 2–ΔCt normalized to the housekeeping ribosomal protein large PO (RPLPO) gene.
Rodari M.M., Cazals-Hatem D., Uzzan M., Martin Silva N., Khiat A., Ta M.C., Lhermitte L., Touzart A., Hanein S., Rouillon C., Joly F., Elmorjani A., Steffann J., Cerf-Bensussan N., Parlato M, & Charbit-Henrion F. (2023). Insights into the expanding intestinal phenotypic spectrum of SOCS1 haploinsufficiency and therapeutic options. Journal of Clinical Immunology, 43(6), 1403-1413.
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6

Quantifying Gene Expression in Tumor Samples

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Total RNA was extracted from fresh tumor tissues, SCC9, and Cal33 cells using RNAeasy Kit (Qiagen) and reversed transcribed. Total RNA was isolated from formalin-fixed, paraffin-embedded tumor tissue sections using RNAeasy FFPE kit (Qiagen) and reverse transcribed. SYBR-Green-1-based RT-PCR amplification was performed in triplicates on the LightCycler-480 (Roche). The primers list is shown in Supplementary Table S2. The relative expression of each gene was analyzed by comparing its expression to that of GAPDH.
Islam S.S., Qassem K., Islam S., Parag R.R., Rahman M.Z., Farhat W.A., Yeger H., Aboussekhra A., Karakas B, & Noman A.S. (2022). Genetic alterations of Keap1 confers chemotherapeutic resistance through functional activation of Nrf2 and Notch pathway in head and neck squamous cell carcinoma. Cell Death & Disease, 13(8), 696.
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7

RNA Extraction and Gene Expression

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409 patients had microarray and clinical data available for analyses. RNA from the primary tumors was extracted by AltheaDx in 2013 according to using Qiagen RNAeasy FFPE kit with a modified deparaffinization step. Gene expression profiles were derived via Affymetrix GeneChip HTA 2.0 (Affymetrix).
Hakimi A.A., Voss M.H., Kuo F., Sanchez A., Liu M., Nixon B., Vuong L., Ostrovnaya I., Chen Y.B., Reuter V.E., Riaz N., Cheng Y., Patel P., Marker M., Reising A., Li M., Chan T.A, & Motzer R.J. (2019). Transcriptomic Profiling of the Tumor Microenvironment Reveals Distinct Subgroups of Clear Cell Renal Cell Cancer - Data from a Randomized Phase III Trial. Cancer discovery, 9(4), 510-525.
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8

Transcriptomic Profiling of Prostate Tumors

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The ECOG-ACRIN biobank retrieved available formalin-fixed, paraffin-embedded (FFPE) biopsy and radical prostatectomy samples from patients enrolled in the CHAARTED trial. De-identified specimens were sent to Decipher Biosciences (San Diego, CA) for central pathology review. The highest grade tumor focus was identified and underwent RNA extraction after macrodissection by a genitourinary pathologist. At least 0.5 mm2 of tumor with at least ≥60% tumor cellularity was required for the assay. RNA was extracted using the RNAeasy FFPE kit (Qiagen, Germantown, MD), converted into cDNA and amplified using the Ovation FFPE kit (TECAN Genomics, Redwood City, CA) and hybridized to the Human Exon 1.0 ST oligonucleotide microarray (ThermoFisher, Carlsbad, CA), as previously described14 (link), in a Clinical Laboratory Improvement Amendments (CLIA)-certified laboratory facility (Decipher Biosciences, San Diego, CA). Quality control was performed using Affymetrix Power Tools, and normalization was performed using the Single Channel Array Normalization (SCAN) algorithm. One hundred and ninety-eight of 790 patients (25%) had banked FFPE tumor blocks available for profiling. Among 190 samples with sufficient tumor available for RNA profiling, a total of 160 samples (84%) passed quality control for downstream analysis.
Hamid A.A., Huang H.C., Wang V., Chen Y.H., Feng F., Den R., Attard G., Van Allen E.M., Tran P.T., Spratt D.E., Dittamore R., Davicioni E., Liu G., DiPaola R., Carducci M.A, & Sweeney C.J. (2021). Transcriptional profiling of primary prostate tumor in metastatic hormone sensitive prostate cancer and association with clinical outcomes: correlative analysis of the E3805 CHAARTED trial. Annals of oncology : official journal of the European Society for Medical Oncology, 32(9), 1157-1166.
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9

Spatial Transcriptomics of FFPE Lung Tissue

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FFPE lung tissue blocks from saline, NTMhigh, BCG, and BCG+NTMhigh groups were soaked in ice-cold water for 2 hours, and 5 μm sections were cut using Leica microtome. Another 10 μm section was subjected to RNA quality assessment using Qiagen RNAeasy FFPE kit and evaluating the DV200 value. Sections were collected in the water bath and allowed to float on the water surface until they were flat and free from wrinkles. Sections were then placed within the frames of capture areas on Visium spatial slides. Slides were then dried at 42°C in an oven and left in a desiccator overnight to ensure complete dehydration.
Dutt T.S., Karger B.R., Fox A., Youssef N., Dadhwal R., Ali M.Z., Patterson J., Creissen E., Rampacci E., Cooper S., Podell B.K., Gonzalez-Juarrero M., Obregon-Henao A, & Henao-Tamayo M. (2022). Mucosal exposure to non-tuberculous mycobacteria elicits B-cell mediated immunity against pulmonary tuberculosis. Cell reports, 41(11), 111783.
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10

Quantification of APOBEC3 Gene Expression

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Primer design for APOBEC3A-G was performed by Primer3 software (https://primer3.ut.ee) after obtaining gene sequences from the UCSC Genome Browser (GRCh38g/hg38) (https://genome.ucsc.edu). Primer pairs were selected so that paired nucleotide sequences remained specific for amplification of individual APOBEC3s and did not overlap with homologous regions of non-target genes, thus eliminating off-target amplification. ACTB was used as reference gene. Primer pairs are shown in Supplementary Table 1.
For gene expression analysis, RNA was extracted from FFPE tissues using the RNAeasy FFPE kit (Qiagen) according to protocol. RNA concentration was measured using the DeNovix fluorescence assay system (DeNovix). For qRT-PCR quantification, at least 250 ng of RNA were used as a template for random hexamer cDNA synthesis using TaqMan Reverse Transcription reagents (ThermoFisher Scientific) according to supplier’s protocol. qRT-PCR was performed with SybrGreen MasterMix (ThermoFisher Scientific) on the QuantStudio7 system (Applied Biosystems). 2 µl of cDNA was used per sample. Melting curve analysis and visualization of PCR products by gel electrophoresis were carried out to verify the presence of specific amplification products.
Trimmel B., Oszwald A., Diemand C., Ertl I.E., Lemberger U., Bruchbacher A., Brettner R., Korn S., Resch I., Comperat E., Shariat S.F, & Hassler M.R. (2022). Evaluation of APOBEC3 expression as prognostic marker in squamous cell carcinoma of the penis. Scientific Reports, 12, 12911.
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