PAXgene Tissue miRNA Kit
For isolation and purification of total RNA, including miRNA, from tissues fixed and stabilized using the PAXgene Tissue Container
- 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
- Reliable quantification of miRNAs
|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)
* PAXgene Tissue fixed
** PAXgene Tissue fixed, paraffin-embedded
The PAXgene Tissue 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 Container or 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.
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 appropriate binding conditions for all RNA molecules 18 nucleotides and longer. The sample is then applied to a PAXgene RNA MinElute spin column, where total RNA binds to the membrane and contaminants are effi ciently washed away. Between the first and second wash steps, the membrane is treated with DNase I to remove trace amounts of bound DNA. After the wash steps, RNA, including miRNA, is eluted in a low-salt elution buffer and denatured by heating.
RNA purification from sections of six different rat PAXgene Tissue fixed, paraffin-embedded (PFPE) tissue types using the PAXgene Tissue RNA Kit or PAXgene Tissue miRNA Kit. Amplification in a quantitative, realtime RT-PCR assay for the miRNA miR-10a using the QIAGEN miScript System.
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) tissue using the PAXgene Tissue miRNA Kit. A scatterplot showing Ct values from different single miRNA-specific RT-qPCR assays using the QIAGEN miScript System: miR9, -10a, -10b, -29a, -103, -125b, -143, -145, -192 and miScript PCR controls RNUA1, RNU5A, RNU6B, SNORD25, SCARNA17, SNORA73A; R2: coefficient of determination.
Total RNA purifed using the PAXgene Tissue 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 Efficient purification of miRNA from PFPE tissue with the PAXgene Tissue miRNA Kit and High concordance of miRNA expression between total RNA isolated from PFPE and flash-frozen tissue).
The PAXgene Tissue 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.
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 miRNA Procedure).
The PAXgene Tissue miRNA Kit provides 3 protocols for purification of RNA, including miRNA, from different starting materials:
- Sections of PFPE tissues
- PF tissue samples (without paraffin embedding)
- Blocks of PFPE tissue samples
Total RNA and miRNA purified with the PAXgene Tissue 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
|miRNA Isolation with the PAXgene Tissue miRNA Kit||957 KB|
|PAXgene Tissue System Brochure||4792 KB|
|Product||Catalog Nr.||Price (USD)|
PAXgene Tissue 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 Containers or PAXgene Tissue FIX Containers + PAXgene Tissue STABILIZER reagent.