PAXgene Tissue STABILIZER

For stabilization of tissue specimens previously fixed in PAXgene Tissue FIX

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PAXgene Tissue STABILIZER Concentrate (150 ml)

Cat. No. / ID: 765512

8 bottles of PAXgene Tissue Stabilizer concentrate, for 4 liters of PAXgene Tissue STABILIZER

For Research Use Only. Not for use in diagnostics procedures. No claim or representation is intended to provide information for the diagnosis, prevention, or treatment of a disease.

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Features

Preservation of both morphology and biomolecules
Tissue can be stored and archived for later processing
RNA, miRNA, DNA, and/or proteins from one sample

Product Details

PAXgene Tissue STABILIZER is used for transport or storage of tissue specimens fixed with the PAXgene Tissue FIX Container (50 ml). After fixation of tissue samples with the PAXgene Tissue FIX Container (50 ml), PAXgene Tissue FIX is removed and replaced by PAXgene Tissue STABILIZER within the same container. Nucleic acids, proteins and morphology of the sample are now stable up to 7 days at room temperature, or for longer periods at 2–8°C, or even at –20°C and –80°C. Stabilized samples can be embedded in paraffin for histological studies. PAXgene Tissue Kits and supplementary protocols provide efficient subsequent purification of RNA, miRNA, DNA, and/or proteins from the same sample.
PAXgene Tissue STABILIZER Concentrate is only to be used in conjunction with tissue specimens previously fixed in PAXgene Tissue FIX.

Performance

The fixation and stabilization method preserves tissue morphology and the integrity of nucleic acids without destructive crosslinking and degradation found in formalin-fixed tissues (see figure " Preservation of tissue morphology").
When fixed tissue is stored in PAXgene Tissue STABILIZER, the nucleic acids, proteins, and morphology of the tissue sample are stable for up to 7 days at room temperature (15–25°C) or for up to 4 weeks at 2–8°C. Tissue samples can even be stored in the PAXgene Tissue STABILIZER for longer periods at –20°C (–15°C to –30°C) or –80°C (–65°C to –90°C) without negative effects on the morphology of the tissue or the integrity of the nucleic acids.
Specifications for storage conditions in PAXgene Tissue STABILIZER were determined using animal tissues.

See figures

Preservation of tissue morphology.

Principle

The methods for tissue fixation currently used in traditional histology are of limited use for molecular analysis. Fixatives that contain formaldehyde crosslink biomolecules and modify nucleic acids and proteins. Such crosslinks lead to nucleic acid degradation during tissue fixation, storage, and processing. Since they cannot be removed completely, the resulting chemical modifications can cause inhibition in downstream applications, such as quantitative PCR or RT-PCR. To enable both molecular and traditional pathology testing from the same specimen, a method is needed to stabilize molecular content and preserve morphology.

To meet this need, PreAnalytiX has developed the PAXgene Tissue System. The system consists of a fixation reagent (PAXgene Tissue FIX), a stabilization reagent (PAXgene Tissue STABILIZER), prefilled containers for tissue collection, storage, and transportation, and kits for purification of RNA, DNA, or total RNA, including miRNA. In addition, supplementary protocols for protein purification and other applications are available in the 'Resources' section of this page or at www.preanalytix.com.

PAXgene Tissue reagents in prefilled containers and PAXgene Tissue Kits provide a complete preanalytical solution for collection, fixation, and stabilization of tissue, and purification of high-quality nucleic acids for molecular research analysis.

Procedure

After fixation of tissue samples with the PAXgene Tissue FIX Container (50 ml), the fixative in the container is removed and replaced by PAXgene Tissue STABILIZER within the same container. Stabilized samples can be embedded in paraffin for histological studies. Nucleic acids and proteins can be isolated from the stabilized samples before or after embedding in paraffin. See the PAXgene Tissue Kit Handbooks for information about DNA, RNA, or miRNA, and the PAXgene Tissue supplementary protocols in the 'Resources' section of this page or at www.preanalytix.com for protein purification and other applications.

Applications

Sections of PAXgene Tissue fixed, paraffin-embedded (PFPE) tissue can be used for histological studies or extraction of nucleic acids or proteins. Purification of RNA, total RNA, including miRNA, or DNA from PAXgene Tissue fixed and stabilized tissue samples requires the use of one of the PAXgene Tissue Kits for RNA, miRNA, or DNA. Purification of protein requires the Qproteome FFPE Kit.

Supporting data and figures

Yield of high-quality, high molecular weight DNA from 12 different PAXgene Tissue-fixed (PF) tissue types, processed on the QIAcube.

Twelve different PAXgene Tissue-fixed (PF) rat tissue types, including fibrous or fatty, DNA-rich and DNA-poor tissues, processed with the PAXgene Tissue DNA Kit on the QIAcube in one single run. DNA yield per 10 mg tissue.

Absorbance ratio of high-quality, high molecular weight DNA from 12 different PAXgene Tissue-fixed, paraffin-embedded (PF) tissue types, processed on the QIAcube.

Comparison of manual and automated procedure: DNA yield from sections of PAXgene Tissue-fixed, paraffin-embedded (PFPE) tissue.

Comparison of manual and automated procedure: Absorbance ratio from sections of PAXgene Tissue-fixed, paraffin-embedded (PFPE) tissue.

DNA without chemical modification can be used for demanding downstream applications.

Protein extracted from PFPE tissue is suitable for two-dimentional gel electrophoresis.

Agarose gel electrophoresis of high-quality, high molecular weight DNA from 12 different PAXgene Tissue-fixed (PF) tissue types, processed on the QIAcube.

Comparison of manual and automated procedure: Agarose gel electrophoresis from sections of PAXgene Tissue-fixed, paraffin-embedded (PFPE) tissue.

Detection of nondegraded, immunoreactive phosphoproteins from human clinical PFPE tissue.

High concordance of miRNA expression between total RNA isolated from PFPE and fresh-frozen tissue.

RNA purified without chemical modification from PFPE tissue using the PAXgene Tissue RNA/miRNA Kit.

Immunohistochemistry (IHC) staining with the PAXgene Tissue System gives comparable results to formalin-fixed tissue.

H&E staining with the PAXgene Tissue System gives results comparable to formalin-fixed tissue.

The PAXgene Tissue FIX Container (50 ml) and PAXgene Tissue STABILIZER workflow.

High-quality DNA from PFPE tissue with preserved morphology.

Resources

Groelz et al., BRN Symposium 2012

Groelz et al., ECP 2012

Groelz et al., BRN Symposium 2011

Groelz et al., 3rd Annual Oncology Biomarkers 2011

Groelz et al., AMP 2009

Groelz et al., ISBER 2012

Groelz et al., AMP 2008

Groelz et al., AACR 2010

Groelz et al., AMP 2010

Groelz et al., AMP 2009

Groelz et al., ECP 2014

Long et al., ADAPT 2012

Hesse et al., AACR-NCI-EORTC 2011

Groelz et al., AMP 2014

For stabilization of tissue specimens fixed in PAXgene Tissue FIX Container

For collection, fixation, and stabilization of multiple small tissue samples or a single large tissue sample

Moving toward excellence and standardization in tissue collection and fixation

For extraction of full-length proteins from PFPE tissue using the Qproteome FFPE Tissue Kit

For isolation and purification of genomic DNA from tissue samples stabilized using the PAXgene Tissue System

Publications

Evaluation of colon cancer histomorphology: a comparison between formalin and PAXgene tissue fixation by an international ring trial.

Gündisch S; Slotta-Huspenina J; Verderio P; Ciniselli CM; Pizzamiglio S; Schott C; Drecoll E; Viertler C; Zatloukal K; Kap M; Riegman P; Esposito I; Specht K; Babaryka G; Asslaber M; Bodó K; den Bakker M; den Hollander J; Fend F; Neumann J; Reu S; Perren A; Langer R; Lugli A; Becker I; Richter T; Kayser G; May AM; Carneiro F; Lopes JM; Sobin L; Höfler H; Becker KF;

Virchows Arch; 2014; 465 (5):509-19 2014 Aug 2 PMID:25085759

Inactivation of Influenza A virus, Adenovirus, and Cytomegalovirus with PAXgene tissue fixative and formalin.

Kap M; Arron GI; Loibner M; Hausleitner A; Siaulyte G; Zatloukal K; Murk JL; Riegman P;

Biopreserv Biobank; 2013; 11 (4):229-34 2013 Aug PMID:24845590

Preservation of nucleic acids and tissue morphology in paraffin-embedded clinical samples: comparison of five molecular fixatives.

Staff S; Kujala P; Karhu R; Rökman A; Ilvesaro J; Kares S; Isola J;

J Clin Pathol; 2013; 66 (9):807-10 2013 Jun 8 PMID:23750036

The Genotype-Tissue Expression (GTEx) project.

GTEx Consortium;

Nat Genet; 2013; 45 (6):580-5 2013 Jun PMID:23715323

The PAXgene(®) tissue system preserves phosphoproteins in human tissue specimens and enables comprehensive protein biomarker research.

Gündisch S; Schott C; Wolff C; Tran K; Beese C; Viertler C; Zatloukal K; Becker KF;

PLoS One; 2013; 8 (3):e60638 2013 Mar 29 PMID:23555997

FAQ

The PAXgene Tissue System involves two processes: fixation and stabilization. PAXgene Tissue FIX provides rapid penetration and fixation that effectively stops all enzymatic activity throughout the tissue. PAXgene Tissue STABILIZER stops fixation and stabilizes the specimen for transportation and storage until processing.

FAQ ID - 3603

Yes. PAXgene Tissue STABILIZER can be used to fill the first position of a tissue processor. See the appendices of the PAXgene Tissue Container Product Circular and PAXgene Tissue FIX Container (50 ml) Product Circular for processing protocols with integrated PAXgene Tissue STABILIZER. When the STABILIZER is included as the first step of a protocol, tissue can be transferred from PAXgene Tissue FIX directly into the first processing position.

FAQ ID - 3608

The PAXgene Tissue System uses an acidic alcoholic fixation without formalin that does not result in cross-linking of biomolecules.

FAQ ID - 3600

No. For storage at –15°C to –30°C or –65°C to –90°C use cryogenic vials with screw caps filled with PAXgene Tissue STABILIZER. For safety reasons, note that the PAXgene Tissue STABILIZER contains 70% ethanol.

FAQ ID -3030

The PAXgene Tissue STABILIZER stabilization reagent contains alcohol and other stabilization agents.

FAQ ID - 3513

Yes, there is co-purification of small RNAs. However, due to the purification chemistry, RNA molecules smaller than approximately 200 nucleotides are recovered with less efficiency. For purification of total RNA, including miRNA and other small RNA molecules of at least 18 nucleotides, use the PAXgene Tissue miRNA Kit.

FAQ ID - 3615

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

FAQ ID - 3613

Parallel processing of formalin-fixed and PAXgene Tissue fixed samples can lead to reduction of nucleic acid yield and integrity from PAXgene Tissue fixed samples through formalin contamination of reagents.

FAQ ID - 3606

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.

FAQ ID - 3610

The stabilization reagent of PAXgene Tissue STABILIZER contains alcohol and other stabilization agents.

FAQ ID - 3602

For samples up to 4 x 15 x 15 mm fixed in a standard tissue cassette, incubate tissue specimen(s) at room temperature (15–25°C) for a minimum of 2 hours, but preferably 3–24 hours. For samples up to 20 x 20 x 20 mm fixed in the PAXgene Tissue FIX Container (50 ml), incubate for 6–48 hours, but preferably 8–24 hours. For biopsies with a thickness of 1 mm or less, fixation time can be reduced to 1 hour. Prolonged fixation times of up to 120 hours (e.g. over the weekend) is possible. Please note that fixation longer than the indicated times may lead to degradation of biomolecules.

FAQ ID -2519

Depending on tissue type, standard storage conditions in PAXgene Tissue STABILIZER are a minimum of 2 hours and up to 7 days at room temperature (15–25°C) or up to 4 weeks at 2–8°C. For longer storage, samples can be kept in PAXgene Tissue STABILIZER at–15°C to –30°C or –65°C to –90°C. Long-term storage studies are ongoing. For latest results, see the poster RNA and Morphology Preservation after 5 years at –20°C and 3 years at –80°C under the Product Resources tab.

FAQ ID - 3605

Yes, human tissue samples treated with the PAXgene Tissue System were successfully used for Fluorescence in situ hybridization (FISH) and Chromogenic in situ hybridization (CISH). However, specimens treated with the PAXgene Tissue System are more sensitive to enzyme digestion compared to formalin-fixed samples. Enzyme digestion and other pretreatment steps may need to be optimized.

FAQ ID -2529

Most antibodies used in IHC assays were developed for use with formalin-fixed tissue and include steps for unmasking epitopes. When working with PAXgene Tissue treated specimens, test for each antibody to determine if it is necessary to perform antigen-retrieval steps. In addition, it may be necessary to optimize antigen-retrieval steps or adjust antibody concentrations in PFPE tissue to achieve optimal staining intensities (see Technical Note Effect of epitope retrieval conditions on immunohistochemical staining of PFPE tonsil tissue with anti-human Ki-67 antigen (clone MIB-1) under the Product Resources tab). Examples for IHC staining of adjacent human tissue sections fixed with neutral-buffered formalin or with PAXgene Tissue reagents are provided in the Tissue Atlas at www.preanalytix.com.

FAQ ID - 3609

Up to 4 standard tissue cassettes, each containing tissue samples with a maximum size of 4 x 15 x 15 mm, can be placed into a PAXgene Tissue FIX Container (50 ml). Alternatively, a single tissue sample with a maximum size of 20 x 20 x 20 mm can be fixed directly in a PAXgene Tissue FIX Container (50 ml). If using a larger tissue sample surrounded by fat (e.g., from a lymph node) or capsule (e.g., from kidney, liver or spleen tissue), partially cut into the tissue every 5 mm (lamination) to enhance permeation of the fixative reagent. If a sample is larger than recommended, the fast and even penetration of fixation reagent is compromised. In such cases, a reduction in the quality of tissue morphology and the integrity of nucleic acids may result.

FAQ ID - 3604

Yes. Supplementary protocols are available for the purification of full-length proteins from PAXgene Tissue fixed (PF) tissue and paraffin blocks using the Qproteome FFPE Tissue Kit (QIAGEN, cat. no. 37623). For more information, see the corresponding supplementary protocols under the Product Resources tab.

FAQ ID - 3616

Human tissue samples treated with the PAXgene Tissue System were successfully used for periodic acid schiff (PAS), resorcin fuchsin, sirius red, and Gomori staining (Kap et al., PLoS ONE 6(11): e27704). However, to achieve the same staining intensities with both PFPE and formalin-fixed, paraffin-embedded (FFPE) samples, it may be necessary to adjust incubation times.

FAQ ID -2530

PFPE tissue blocks can be stored at 2–25°C for short-term storage or transport. However, biomolecules within paraffin blocks will undergo slow chemical degradation. To better preserve morphology and biomolecule integrity within the paraffin-embedded tissue, store PFPE blocks at–30°C to –15°C. For the latest results on long-term storage of PFPE and formalin-fixed, paraffin-embedded (FFPE) tissue, see poster RT-PCR Performance of RNA Obtained from Archived FFPE and PFPE Blocks of Tissue under the Product Resources tab.

FAQ ID -2524

Yes. Comparable morphology was observed in adjacent pieces from a tissue sample fixed either with neutral-buffered formalin or with the PAXgene Tissue System for a variety of human and animal tissue (Gündisch et al., Virchows Arch. 2014 Aug; Kap et al., PLoS ONE 6(11): e27704). Examples are provided in the Tissue Atlas at www.preanalytix.com. PAXgene Tissue treated specimens have a tendency to be more eosinophilic. If an identical staining pattern to formalin-fixed samples is required, the incubation time in eosin should be reduced.

FAQ ID -2526

Yes. PreAnalytiX has developed a workflow and protocols for cryo-embedding tissue specimens fixed and stabilized in either the PAXgene Tissue Container or the PAXgene Tissue FIX Container (50 ml). A supplementary protocol for generating PAXgene® Tissue fixed, cryo-embedded (PFCE) tissues is available under the Product Resources tab.

FAQ ID -2525

RNA yield depends on several parameters, such as tissue type, time from resection until fixation, fixation time (ideally 2 to 4 hours), 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 Product Resources tab).

FAQ ID -2534

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

FAQ ID - 3614

The PAXgene Tissue fixation and stabilization chemistry is free of cross-linking reagents. RNA 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).

FAQ ID -2538

PAXgene Tissue Kits can be used for extraction 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 Product Resources tab.

FAQ ID - 3617

The PAXgene Tissue DNA, RNA, and 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 and miRNA purified with the PAXgene Tissue RNA or miRNA Kits are both >1.8.

For examples of RNA purity see Technical Note "Yield, purity, and integrity of RNA purified from PAXgene Tissue fixed, paraffin-embedded (PFPE) rat tissue" under the Product Resources tab.

FAQ ID -2533

The fixation reagent of PAXgene Tissue FIX contains alcohols and an acid, in addition to other stabilization agents.

FAQ ID - 3601

Yes. All processors commonly used for formalin-fixed samples can be used to produce PAXgene Tissue fixed, paraffin-embedded (PFPE) blocks of tissue. However, when processing PAXgene Tissue-treated specimens, do not use reagents contaminated with formalin. Residual formalin can lead to significant reduction of nucleic acid yield and integrity from PFPE tissue samples (see Technical Note “Influence of formalin contamination during processing of PAXgene Tissue fixed, paraffin-embedded tissue (PFPE) on RNA yield, integrity, and performance in quantitative RT-PCR”). Therefore, always use separate batches of reagents (alcohol, xylene, or xylene substitutes) for processing PAXgene Tissue fixed and formalin-fixed samples.

FAQ ID -3032

In contrast to DNA isolated from formalin-fixed, paraffin-embedded (FFPE) tissue, DNA from PFPE tissue exhibits high molecular weight. In most cases, a distinct 10 kb band is observed in electrophoretically separated DNA eluates. For an example, see Figure 2 in the Technical Note Quantitative analysis of KRAS and BRAF mutational status in DNA from PAXgene Tissue fixed, paraffin-embedded (PFPE) tissue using Pyrosequencing technology under the Product Resources tab.

FAQ ID - 3612

Yes. Supplementary protocols for generating sections from PAXgene Tissue fixed, Paraffin-embedded (PFPE) and PAXgene Tissue fixed, cryo-embedded (PFCE) tissue blocks for manual and laser microdissection are available under the Product Resources tab.

FAQ ID -2531

Similar to yield, RNA integrity depends on several parameters, such as tissue type, time from resection until fixation, fixation time, processing protocol, 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).

FAQ ID - 3611

Proteins from PAXgene Tissue fixed, PAXgene Tissue fixed, paraffin-embedded (PFPE) and PAXgene Tissue fixed, cryo-embedded (PFCE) tissues are non-degraded, immunoreactive and have been successfully investigated by western blot analysis, reverse-phase protein arrays, two-dimensional gel electrophoresis (2D-PAGE), enzyme-linked immunosorbent assay (ELISA),and MALDI imaging mass spectrometry.

FAQ ID - 3619

In contrast to FFPE, DNA purified from PFPE and PFCE is of high molecular weight, free of chemical modifications. In demanding downstream applications, such as multiplex or long-range PCR, it performs similarly or identically to DNA isolated from frozen tissue. For examples, see, Figure 5 in Viertler et al., J Mol Diagn. 2012 Sep; 14(5).

FAQ ID - 3618

Fixation is performed within a standard tissue cassette. The maximum sample size is 4 x 15 x 15 mm, so that the sample fits in the cassette without requiring physical pressure to close the lid. The cassette should leave no marks or grid impressions on the tissue.

FAQ ID -2518

No. Special cleaning is not required. However, be sure to change the alcohol and xylene (or xylene substitute), and keep separate batches of reagents for processing PAXgene Tissue fixed samples. Reagents contaminated with formalin can lead to reduced nucleic acid yields and integrity.

FAQ ID - 3607