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PAXgene Tissue RNA/miRNA Kit

For isolation and purification of total RNA, including miRNA, from tissues fixed and stabilized using the PAXgene Tissue System

  • Effective purification of high and low molecular weight RNA from tissue with preserved morphology
  • Protocols for paraffin-embedded or paraffin-free tissues
  • Minimal genomic DNA contamination

 

Feature Specification
Format Spin column
Technology Silica membrane
Main sample type Human tissue
Sample amount Up to 10 mg from PF* tissue
2–5 sections 10 µm thick from PFPE** tissue with a maximum
size of 15 x 15 mm
Elution volume 14–40 µl
Time per run 70 min + 15 min incubation/8 samples
(120 min for fibrous tissues)
Processing Manual: centrifugation

* PAXgene Tissue fixed
** PAXgene Tissue fixed, paraffin-embedded

 

Intended Use

The PAXgene Tissue RNA/miRNA Kit is intended for the purification of total RNA, including miRNA, from PAXgene Tissue fixed (PF) and PAXgene Tissue fixed, paraffin-embedded (PFPE) tissue samples. The kit is intended to be used as part of the PAXgene Tissue System, which additionally includes the PAXgene Tissue FIX Container and the PAXgene Tissue STABILIZER Concentrate for the collection, fixation, stabilization, and storage of tissue samples.

For research use only. Not for use in diagnostic procedures. No claim or representation is intended to provide information for the diagnosis, prevention, or treatment of a disease. The performance characteristics of this product have not been fully established.

The PAXgene Tissue RNA/miRNA Procedure
Disruption and homogenization of the tissue sample is performed in binding buffer, Buffer TM1. After centrifugation to remove residual cell debris, isopropanol is added to the lysate to provide [...]
High concordance of miRNA expression between total RNA isolated from PFPE and flash-frozen tissue
RNA including miRNA purified from mirrored human breast cancer tissue fresh frozen in liquid nitrogen using the QIAGEN miRNeasy Kit, or from sections of PAXgene Tissue fixed, paraffin-embedded (PFPE) [...]
RNA purified without chemical modification from PFPE tissue using the PAXgene Tissue RNA/miRNA Kit
SYBR Green real-time RT-qPCR was performed with 10 ng RNA from cryopreserved, formalin-fixed, paraffin-embedded (FFPE) or PAXgene Tissue-fixed, paraffin-embedded (PFPE) rat tissue (modified according [...]
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Performance

Total RNA purifed using the PAXgene Tissue RNA/miRNA Kit is highly pure. Genomic DNA contamination is minimized and the purified RNA is ready to use in downstream applications with no detectable inhibition of PCR. The resulting eluate includes smaller RNA molecules, such as 5.8S rRNA, 5S rRNA, tRNAs, and miRNAs. Studies have shown that miRNA purified from PFPE tissue samples is reliably quantified, with high concordance to flash-frozen samples (see High concordance of miRNA expression between total RNA isolated from PFPE and flash-frozen tissue).

 

Principle

The PAXgene Tissue RNA/miRNA Kit enables purification of total RNA from tissues fixed and stabilized using the PAXgene Fixation and Stabilization products, which preserve tissue morphology and biomolecule integrity by avoiding destructive cross-linking and degradation found in formalin-fixed tissues. The purified RNA and miRNA have no inhibitory chemical modifications and thus, can be used in sensitive downstream applications.

 

Procedure

PAxgene Tissue fixed (PF) or PAXgene Tissue fixed, paraffin-embedded (PFPE) tissue samples are disrupted and homogenized in binding buffer. After centrifugation to remove cellular debris, optimal conditions are created for binding of RNA molecules to the silica membrane. Contaminants are then washed away and DNase I treatment removes any trace amounts of bound DNA. Total RNA, including miRNA, is then eluted in a low-salt elution buffer and denatured by heating (see The PAXgene Tissue RNA/miRNA Procedure).

 

The PAXgene Tissue RNA/miRNA Kit provides 2 protocols for purification of RNA, including miRNA, from different starting materials:

  • Sections of PFPE tissues
  • PF tissue samples (without paraffin embedding)

 

Applications

Total RNA and miRNA purified with the PAXgene Tissue RNA/miRNA Kit is ready to use in a range of downstream research applications (see Resources), including:

  • cDNA synthesis
  • Gene expression arrays
  • End-point RT-PCR
  • Quantitative RT-PCR
  • Detection and quantification of miRNA
  • RNA sequencing
 

Handbooks (1)

File sizeFormatDownload
PAXgene Tissue miRNA Handbook 381 KB pdf

Supplementary Protocols (4)

File sizeFormatDownload
Purification of total RNA, including miRNA, from sections of PAXgene Tissue fixed, cryo-embedded (PFCE) tissue placed directly into a microcentrifuge tube 621 KB pdf
Simultaneous purification of genomic DNA and total RNA, including miRNA, from sections of PAXgene Tissue fixed, paraffin-embedded (PFPE) tissue 85 KB pdf
Deparaffinization of PAXgene Tissue fixed, paraffin-embedded tissue sections with Deparaffinization Solution 86 KB pdf
Purification of total RNA, including miRNA, from microdissected PAXgene® Tissue fixed, paraffin embedded (PFPE) and PAXgene Tissue fixed, cryoembedded (PFCE) tissues 679 KB pdf

Brochures (2)

File sizeFormatDownload
miRNA Isolation with the PAXgene Tissue miRNA Kit 957 KB pdf
PAXgene Tissue System Brochure 4792 KB pdf

Technical information (2)

File sizeFormatDownload
Influence on RNA yield and integrity of modifications to the processing protocol 648 KB pdf
Influence of formalin contamination during processing of PAXgene Tissue fixed, paraffin-embedded tissue (PFPE) on RNA yield, integrity, and performance in quantitative RT-PCR 677 KB pdf

Safety Data Sheets (1)

File sizeFormatDownload
MSDS PAXgene Tissue miRNA Kit Link

Posters (4)

File sizeFormatDownload
A New Tissue Fixative for Biomarker Discovery: Gene expression and miRNA in PAXgene® Tissue Fixed Paraffin-Embedded (PFPE) Colorectal Cancer (CRC) and Breast Cancer Tissue vs. Formalin Fixed Paraffin-Embedded (FFPE) Tissue; Groelz et al., 3rd Annual Oncology Biomarkers 2011 423 KB pdf
PAXgene Tissue: A New Fixation Technology for Simultaneous Preservation of Morphology and Nucleic Acids; Groelz et al., BRN Symposium 2011 555 KB pdf
Evaluation of the PAXgene® Tissue System: Preservation of morphology and gene expression in human melanoma; Hesse et al., AACR-NCI-EORTC 2011 997 KB pdf
PAXgene Tissue Fixation Technology for Simultaneous Preservation of Morphology and Biomolecules; Groelz et al., ECP 2012 1231 KB pdf

References (9)

File sizeFormatDownload
Feature-based analysis of mouse prostatic intraepithelial neoplasia in histological tissue sections. Ruusuvuori P. et al., J Pathol Inform. 2016 Jan 29;7:5 Link
Pathogen Inactivating Properties and Increased Sensitivity in Molecular Diagnostics by PAXgene, a Novel Non-Crosslinking Tissue Fixative. Loibner M. et al., PLoS One. 2016 Mar 14;11(3):e0151383. Link
Protocol for HER2 FISH determination on PAXgene-fixed and paraffin-embedded tissue in breast cancer. Oberauner-Wappis L et al., Int J Exp Pathol. 2016 Apr;97(2):202-6 Link
The focus on sample quality: Influence of colon tissue collection on reliability of qPCR data. Korenkova V et al., Sci Rep. 2016 Jul 7;6:29023. doi: 10.1038/srep29023 Link
A Critical Evaluation of the PAXgene Tissue Fixation System:  Morphology, Immunohistochemistry, Molecular Biology, and Proteomics. Mathieson W. et al., Am J Clin Pathol. 2016 Jul;146(1):25-40. Link
Assessment of PAXgene Fixation on Preservation of Morphology and Nucleic Acids in Microdissected Retina Tissue. Liu Y and Edward DP, Curr Eye Res. 2016 Jul 13:1-7. Link
Surgical Specimens of Colorectal Cancer Fixed with PAXgene Tissue System Preserve High-Quality RNA; Hara K. et al., Biopreserv Biobank. 2015 Oct;13(5):325-34 Link
A Novel Approach to High-Quality Postmortem Tissue Procurement: The GTEx Project; Carithers et al., Biopreservation and Biobanking. October 2015, 13(5): 311-319 931 KB pdf
Next-gen tissue: preservation of molecular and morphological fidelity in prostate tissue; Gillard M. et al., Am J Transl Res 2015;7(7):1227-1235 Link

FAQ (8)

    Purification and quality of biomolecules from PAXgene Tissue-treated samples

    1. Is it possible to use formalin-fixed, paraffin-embedded (FFPE) kits and protocols to isolate biomolecules from PAXgene Tissue fixed, paraffin-embedded (PFPE) tissues?

    No. Procedures developed for the extraction of biomolecules from FFPE tissues include prolonged proteinase K digestion and heating steps to remove chemical modifications introduced by formalin. Since the PAXgene Tissue System does not chemically modify biomolecules, these steps are not necessary and, in fact, lead to degradation of biomolecules. Instead, use dedicated PAXgene Tissue Kits and supplementary protocols for extraction of biomolecules from PAXgene Tissue-treated samples.

    2. What is the purity of nucleic acids extracted with the PAXgene Tissue Kits?

    The PAXgene Tissue DNA and RNA, including miRNA, Kits are based on proven QIAGEN technologies. Nucleic acids isolated with these kits are generally of high purity.

    On average, measurements of the A260/A280 ratio for DNA purified with the PAXgene Tissue DNA Kit are >1.7, and ratios for RNA including miRNA purified with the PAXgene Tissue RNA/miRNA Kit are >1.8.

    3. What is the average RNA yield, including miRNA, from PAXgene Tissue fixed, paraffin-embedded (PFPE) tissues?

    RNA yield, including miRNA, depends on several parameters, such as tissue type, time from resection until fixation, fixation time, processing protocol used and age and storage conditions of the PFPE block.

    In a study with PFPE tissue sections (area: 100 mm²;  thickness: 10 µm) median RNA yield from rat liver was 4.2 µg (n=58), from kidney 2.2 µg (n=58), from spleen 4.7 µg (n=58), from intestine 4.7 µg (n=58) and from lung 0.9 µg (n=58). See Technical Note "Yield, purity, and integrity of RNA purified from PAXgene Tissue fixed, paraffin-embedded (PFPE) rat tissue" under the Resources tab.

    4. How well is RNA integrity preserved in PAXgene Tissue fixed, paraffin-embedded (PFPE) tissues?

    Similar to yield, RNA integrity depends on several parameters, such as tissue type, time from resection until fixation, fixation time, processing protocol and age and storage conditions of the PFPE block. For examples of RNA integrity values from rat tissues under ideal workflow conditions, see Groelz et al., Exp Mol Pathol. 2013 Feb; 94(1). For examples of RNA integrity from clinical samples, see Viertler et al., J Mol Diagn. 2012 Sep; 14(5).

    5. What are the yield and integrity of nucleic acids isolated from blocks of PAXgene Tissue fixed, cryo-embedded (PFCE) tissues?

    DNA and RNA isolated from PFCE tissue specimens are of high quantity and quality, comparable to PFPE tissue.

    6. Are special kits and protocols required for the isolation of biomolecules from PAXgene Tissue fixed, cryo-embedded (PFCE) tissues?

    No. Regular PAXgene Tissue kits can be used for the isolation of RNA, miRNA and DNA from PFCE tissue. Supplementary protocols developed specifically for the extraction of biomolecules from PFCE samples are available under the Resources tab.

    7. Which kits and protocols can be used for the isolation of nucleic acids from microdissected PAXgene Tissue fixed, paraffin-embedded (PFPE) and PAXgene Tissue fixed, cryo-embedded (PFCE) specimens?

    Regular PAXgene Tissue kits can be used for the isolation of RNA, miRNA and DNA from microdissected PFPE and PFCE tissue. Supplementary protocols developed specifically for the extraction of biomolecules from microdissected PFPE and PFCE samples are available under the Resources tab.

    Molecular analysis of biomolecules purified from PAXgene Tissue-treated samples

    1. What is the RT-PCR performance of RNA purified from PAXgene Tissue fixed, paraffin-embedded (PFPE) and PAXgene Tissue fixed, cryo-embedded (PFCE) tissues compared to RNA from snap frozen or formalin-fixed, paraffin-embedded (FFPE) tissues?

    RNA including miRNA purified from PFPE and PFCE is free of chemical modifications and performs similarly or identically to RNA isolated from frozen tissue. For examples of the correlation of gene expression levels in snap frozen tissue, FFPE and PFPE, see Figure 4 in Groelz et al., Exp Mol Pathol. 2013 Feb; 94(1) and Figure 3 in Viertler et al., J Mol Diagn. 2012 Sep;14(5).

     

ProductCatalog Nr.Price (USD)

PAXgene Tissue RNA/miRNA Kit (50)

For 50 RNA preps: PAXgene RNA MinElute Spin Columns, PAXgene Shredder Spin Columns, Processing Tubes, Microcentrifuge Tubes, Carrier RNA, RNase-Free DNase, and RNase-Free Buffers. To be used in conjunction with PAXgene Tissue FIX Containers + PAXgene Tissue STABILIZER reagent.
766134640.00