RNAprotect Reagents and Tubes

For immediate stabilization of RNA in tissues, cultured cells and bacterial cultures

Features

  • Immediate stabilization of RNA and RNA protection
  • Convenient and safe handling at room temperature
  • Easy collection and storage of tissues in reclosable tubes
  • Stabilization of Gram-positive and Gram-negative bacteria
  • No need to remove medium from sorted or cultured cells
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RNAprotect Tissue Reagent (50 ml)

Cat. No. / ID: 76104

50 ml RNAprotect Tissue Reagent for stabilization of RNA in 25 x 200 mg tissue samples
ReagentTube
RNAprotect Reagent
RNAprotect Tubes
Sample type
Tissue
Bacteria
Cells
Quantity
50 ml
250 ml
RNAprotect Reagents and Tubes are intended for molecular biology applications. These products are not intended for the diagnosis, prevention, or treatment of a disease.

Product Details

RNAprotect Reagents and Tubes provide immediate RNase inactivation and stabilization of RNA in tissues, cultures of Gram-positive and Gram-negative bacteria and sorted or cultured cells to preserve the gene expression profile and enable reproducible purification of high-quality RNA. Samples are stabilized at room temperature prior to RNA isolation procedures.

Performance

RNAprotect Tissue Reagent ensures immediate RNase inactivation and stabilizes and protects the RNA expression pattern of samples during harvest and storage (see figure " RNAprotect Reagent prevents degradation of mRNA in tissues"). Immediate protection with RNAprotect technology ensures that downstream analyses truly reflect the expression profile of the intact tissues. Samples can be archived without risk of RNA degradation, even after multiple freeze–thaw cycles (see figures “ Stable RNA in tissues: different temperatures and freeze–thaw cycles” and “ RNAprotect Tissue Tubes prevent RNA changes in tissues”).


RNAprotect Cell Reagent prevents induction of gene expression in treated cells and preserves and stabilizes transcript levels during storage of cell samples (see figures " Effective inhibition of PMA induction of c-fos expression" and " More reliable quantification of CD83 transcript").


RNAprotect Bacteria Reagent is suitable for use with a wide range of bacterial species, both Gram positive (e.g., Staphylococcus aureus and Mycobacterium avium) and Gram negative (e.g., Escherichia coli and Salmonella typhimurium) and prevents RNA changes (see figures “ RNAprotect Bacteria Reagent prevents mRNA degradation” and “ GeneChip analysis”). Bacteria can be grown in either minimal or complex medium prior to isolation of RNA.

GeneChip analysis.
RNeasy purified RNA from RNAprotect stabilized samples stored under a wide variety of conditions is ideal for downstream processes such as northern blotting and RT-PCR.

See figures

Principle

Once a biological sample is harvested, its RNA becomes extremely unstable. RNAprotect Reagents and Tubes enable immediate RNase inactivation, stabilization of the RNA and preservation of the RNA expression pattern. Gene induction or down-regulation triggered by sample manipulation is also prevented, allowing accurate analysis of gene expression. This is particularly important for bacterial expression profiles. Enzymatic degradation of RNA leads to reduction or loss of many transcripts because bacterial mRNA molecules have very short half lives of only a few minutes.


RNAprotect Tissue Reagent quickly permeates tissues resulting in immediate stabilization of the RNA. RNAprotect Cell Reagent provides immediate stabilization of RNA in sorted or cultured cells. Cells are stabilized at room temperature and can be stored or transported at ambient temperature prior to RNA purification.

Procedure

For RNA protection and sample storage, tissue samples are submerged in an appropriate volume of RNAprotect Tissue Reagent immediately after harvesting. RNAprotect Tissue Tubes provide premeasured volumes of RNAprotect Tissue Reagent in reclosable tubes for convenient handling and sample storage. The reagent preserves RNA for up to 1 day at 37°C, 7 days at 18–25°C, or 4 weeks at 2–8°C, allowing processing, transportation, storage, and shipping of samples without liquid nitrogen or dry ice. Alternatively, the samples can also be placed at –20°C or –80°C for archival storage.


RNAprotect Cell Reagent immediately stabilizes the gene expression profile in cells. With other technologies, it is necessary to wash or process the cells (e.g., trypsin treatment) prior to adding an RNA stabilization reagent. Changes in expression that may occur during the washing or processing steps are eliminated by adding RNAprotect Cell Reagent directly to cells in culture medium or buffer. Instantaneous RNA stabilization with RNAprotect Cell Reagent can also be extended to pelleted cells not in medium or storage solution. The amount of starting material must not exceed 1 x 107 cells.


For bacterial samples, two volumes of RNAprotect Bacteria Reagent are added directly to 1 volume of bacterial culture prior to RNA isolation, providing immediate stabilization of RNA. The stabilization allows time for efficient bacterial lysis using a choice of protocols: enzymatic lysis, mechanical disruption, or a combination of both methods. QIAGEN recommends the TissueLyser for efficient mechanical disruption.

Applications

RNA stabilized in RNAprotect Reagents and Tubes can be used for:

  • Total RNA purification
  • miRNA purification
  • miRNA purification

Supporting data and figures

Resources

Kit Handbooks (3)
RNAprotect Tissue Tubes - For collection of harvested animal tissues with immediate stabilization of the gene expression profile, and subsequent transport and storage; RNAprotect Tissue Reagent - For immediate stabilization of the gene expression profile in harvested animal tissues

Publications

Dll4 signalling through Notch1 regulates formation of tip cells during angiogenesis.
Hellström M; Phng LK; Hofmann JJ; Wallgard E; Coultas L; Lindblom P; Alva J; Nilsson AK; Karlsson L; Gaiano N; Yoon K; Rossant J; Iruela-Arispe ML; Kalén M; Gerhardt H; Betsholtz C;
Nature; 2007; 445 (7129):776-80 2007 Jan 28 PMID:17259973
Gene networks and microRNAs implicated in aggressive prostate cancer.
Wang L; Tang H; Thayanithy V; Subramanian S; Oberg AL; Cunningham JM; Cerhan JR; Steer CJ; Thibodeau SN;
Cancer Res; 2009; 69 (24):9490-7 2009 Dec 15 PMID:19996289
Determinants of RNA quality from FFPE samples.
von Ahlfen S; Missel A; Bendrat K; Schlumpberger M;
PLoS One; 2007; 2 (12):e1261 2007 Dec 5 PMID:18060057
Novel retinoid targets in the mouse limb during organogenesis.
Ali-Khan SE; Hales BF;
Toxicol Sci; 2006; 94 (1):139-52 2006 Jun 13 PMID:16772331
Characterization of Bacillus anthracis-like bacteria isolated from wild great apes from Cote d'Ivoire and Cameroon.
Klee SR; Ozel M; Appel B; Boesch C; Ellerbrok H; Jacob D; Holland G; Leendertz FH; Pauli G; Grunow R; Nattermann H;
J Bacteriol; 2006; 188 (15):5333-44 2006 Aug PMID:16855222

FAQ

Can you recommend a Reagent or Kit for stabilizing RNA in cultured cells?

Yes, we recommend to use the RNeasy Protect Cell Mini Kit which contains RNAprotect Cell Reagent, the first RNA stabilization reagent for cultured cells in the market optimized for a broad range of cell types and cell culture formats.

 

FAQ ID -1215
How long can I store bacterial culture stabilized in RNAprotect Bacteria Reagent?
Bacterial cultures stabilized in RNAprotect Bacteria Reagent can be stored at room temperature for up to 3 hours. Alternatively, pellets obtained after centrifugation of the bacterial culture/RNAprotect Bacteria Reagent mixture can be frozen and stored at -20ºC for up to 2 weeks, or at -70ºC for up to 4 weeks.
FAQ ID -614
Is RNAprotect Bacteria Reagent compatible with the RNeasy Mini Kit?

Yes, RNAprotect Bacteria Reagent and the RNeasy Mini Kit are compatible. You can also purchase QIAGEN's RNeasy Protect Bacteria Kits, which will contain RNAprotect Bacteria Reagent and RNeasy chemistry. Protocols for isolating RNA from bacteria after using the Reagent for bacterial RNA stabilization can be found in the RNAprotect Bacteria Reagent Handbook.

FAQ ID -797
Do you have a kit for RNA isolation from any kind of sample type?

The RNeasy 96 Universal Tissue Kit enables high-throughput purification of RNA from any animal or human tissue sample, including difficult-to-lyse fibrous and fatty tissues. For single tube format RNA purification, the RNeasy Plus Universal Kit is also available. 

Please refer to the Selection guide for RNA isolation for all sample types to find the optimal solution for your sample source.

FAQ ID -627
Do you have a protocol for the isolation of RNA from bacterial cultures using RNAprotect Bacteria Reagent and QIAzol Lysis Reagent?
Yes, please follow the protocol in the RNAprotect Bacteria Reagent Handbook in Appendix D:  Stabilization and Purification of Bacterial RNA Using RNAprotect Bacteria Reagent, the RNeasy Lipid Tissue Mini Kit, and the TissueLyser.
FAQ ID -1020
Why is it not recommended to stabilize cells with RNAprotect Tissue Reagent?

RNAprotect Tissue Reagent is recommended for stabilization and protection of RNA in animal tissues. It is not recommended for cultured cells because it may be difficult to pellet and remove them from the reagent prior to RNA isolation (see FAQ 364). 

Instead, we recommend to use the RNAprotect Cell Reagent.  This reagent is for immediate RNA stabilization in sorted or cultured cells with no need to remove medium

 

FAQ ID -941
Is it possible to perform laser microdissection with tissues stabilized with RNAprotect Tissue reagent?

For Laser Captured Microdissection, its best to either flash-freeze the tissue and do cryosectioning, or to use FFPE material. 

FAQ ID -608
Can I stabilize adipose tissue using RNAprotect Tissue Reagent prior to RNA isolation?

In general, this is possible and has been done successfully in our labs with brain tissue. However, if the starting material for RNA isolation is extremely fatty, it will be difficult for the aqueous RNAprotect Tissue Reagent to permeate into the tissue and therefore, to sufficiently stabilize it.


FAQ ID -856
Is RNAprotect Cell Reagent provided in the RNeasy Protect Cell Kit available separately?

Yes, RNAprotect Cell Reagent of the RNeasy Protect Cell Mini Kit can be purchased separately in a bulk size of 250 ml for processing large cell culture volumes.

 

FAQ ID -1217
What is the recommended incubation time to stabilize tissue RNA in RNAprotect Tissue Reagent?

The RNAprotect Tissue Reagent penetrates the sample by diffusion and protects RNA immediately upon contact with the surface layer and outer portions of solid tissues. In order to ensure reliable stabilization of RNA even in the inner parts of solid tissues, we recommend incubation for at least 45 minutes in the reagent prior to RNA isolation using RNeasy Kits. Note that tissue size is critical for successful RNA stabilization with RNAprotect technology. Samples must be cut into slices less than 0.5 cm thick for rapid and reliable stabilization in interior tissue parts. RNA degradation will occur in tissue slices that are too thick.

For archival storage in the freezer, first incubate the sample overnight in the reagent at 2–8°C. Please see detailed instructions for storage under step 5 of the “Protocol: Stabilization of RNA in Harvested Animal Tissues” in the RNAprotect Handbook.


FAQ ID -647
Can RNAprotect Tissue Reagent be used to stabilize tissue which has already been flash-frozen?

No, we do not recommend to use the RNAprotect Tissue Reagent to stabilize flash-frozen tissue. It should only be used to stabilize fresh tissue.


FAQ ID -757
Do you have a protocol for the preparation of tissues preserved in RNAprotect Tissue Reagent for pathological studies?

Yes, please follow the User-Developed Protocol 'Preparation of RNAprotect preserved tissues for pathological studies' (PROM-15329).


FAQ ID -950
Can RNAprotect Tissue Reagent be used to store previously isolated RNA?

We do not recommend to store previously isolated RNA in RNAprotect Tissue Reagent, since it will be very difficult to recover isolated RNA from it. Purified RNA can be stored at −30 to −15°C or −90 to −65°C in sterile RNase-free water. Under these conditions, we found no detectable RNA degradation after 1 year of storage.


FAQ ID -337
How much RNA does a bacterial cell contain?
The RNA content and make up of a bacterial cell is highly dependent on the type of bacteria, and the developmental and physiological state of the cell. To estimate the approximate yield that can be expected from your starting material, we usually calculate that a typical bacterial cell contains 100 fg of total RNA.
FAQ ID -2949
Why are some of my RNAprotect-stabilized samples frozen at 0ºC while others are not?

Different samples of varying water content may affect the point at which the reagent will form a solid at low temperature. Other factors can be temperature variation from freezer to freezer, or even within the same freezer (e.g., different temperatures near the door and in the back region of the freezer). Regardless, the performance of the RNAprotect Tissue Reagent and the RNA quality are not affected by freezing of the reagent.


FAQ ID -758
Can I stabilize RNA in bacteria grown on solid media or substrate using the RNAprotect Bacteria Reagent?

Yes, but you have to make sure that the RNAprotect Bacteria Reagent is diluted in PBS or water prior to use (2 volumes reagent : 1 volume PBS or water) in order to achieve its optimal working concentration. The diluted reagent is directly added to the bacteria growing on the solid substrate, and reagent and bacteria are mixed thoroughly. The reagent is not recommended for stab cultures, since sufficient access of the reagent to all bacteria for immediate RNA stabilization cannot be guaranteed.

FAQ ID -615
What has to be done to an RNA sample before loading it onto an Agilent Bioanalyzer?

For RNA isolated on the BioRobot EZ1 and BioRobot M48:

The RNA can be directly applied to the Agilent Bioanalyzer, since it is being denatured in the final protocol steps of these isolation procedures.

For RNA prepared with all other QIAGEN RNA Isolation Products:

We recommend to denature the samples in a water bath for 2 min at 70°C, and then place them directly on ice prior to loading them onto the Agilent Bioanalyzer.

 

FAQ ID -528
I accidentally added RNAprotect Tissue Reagent to my cells, and now the cells are difficult to pellet. What can I do?

If the cells in RNAprotect Tissue Reagent cannot be collected by centrifugation, please try one of the following suggestions:

1) Add 600 µl Buffer RLT to a maximum of 200 µl sample volume, and proceed with step 3 of the "RNeasy Mini Protocol for Isolation of Total RNA from Animal Cells" in the 
RNeasy Mini Handbook. Load the lysate onto the column in successive aliquots in step 5 of the protocol.

2) If cells are floating on the surface of the RNAprotect Tissue Reagent, try removing the reagent by pipetting from underneath. Leave behind approximately 100 ul of RNAprotect Tissue Reagent, and add 350 ul Buffer RLT before proceeding with the protocol "RNeasy Mini Protocol for Isolation of Total RNA from Animal Cells". For every 100 ul of cells in RNAprotect Tissue Reagent, use 250 ul of 96-100% ethanol instead of the 70% ethanol listed in step 4 of the standard protocol.

3) Dilute the sample 10x by adding cold PBS. Pellet cells by centrifugation. Caution: Cells might lyse.

4) Try to pipette the floating cells off the surface.

Note that the above steps are suggestions, rather than official protocol recommendations. Please try a "pilot" run on a test sample first.

For RNA stabilization of cells, we recommend using the RNAprotect Cell Reagent.

 

FAQ ID -364
How long can I store tissues preserved in RNAprotect Tissue Reagent?

Tissues preserved in RNAprotect Tissue Reagent can be stored as long as listed below:

Temp [°C] Duration
37 °C 1 day
room temp. 1 week
2-8 °C 1 month
-30 to -15 °C archival
-90 to -65 °C archival
FAQ ID-3750
Will pancreas tissue swim on top of RNAprotect Tissue Reagent?

We never have seen it in-house for mouse or rat pancreas, but if a lot of fatty tissue is present in the tissue sample it may float on top. Our R&D department has obtained good results with RNAprotect Tissue Reagent preserved mouse and rat pancreas.


FAQ ID -536
Can cells stabilized with RNAprotect Cell Reagent be used for cell sorting by flow cytometry (FACS)?

We do not recommend to use cells stabilized with RNAprotect Cell Reagent for cell sorting as cell morphology will be altered after adding the reagent, and stabilized cells may lyse during the sorting process.

 

FAQ ID -1216