Urine microRNA Purification Kit
Rapid purification of microRNA from urine without phenol
For research use only and NOT intended for in vitro diagnostics.
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Rapid purification of microRNA from urine without phenol
$279.00
SKU
29000
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Overview
- Isolate high quality total RNA (including small RNA and microRNA) and all sizes of circulating and exosomal RNA, including microRNA from urine and cerebrospinal fluid (CSF) samples
- Small urine and CSF input ranging from as low as 0.5 mL to 1 mL
- No phenol extractions
- Very sensitive and linear down to a few cells without the need for carrier RNA
- Bind and elute all RNA irrespective of size or GC content, without bias
- Rapid and convenient spin column protocol
- Purification is based on spin column chromatography that uses Norgen’s proprietary resin separation matrix
Norgen’s Urine microRNA Purification Kit provides a rapid method for the isolation and purification of small RNA molecules (All sizes, including < 200 nt) from urine samples. These small RNAs include regulatory RNA molecules such as microRNA (miRNA) as well as tRNA and 5S rRNA. Small RNA molecules are often studied due to their ability to regulate gene expression. Typically miRNAs are 20-25 nucleotides long and regulate gene expression by binding to mRNA molecules and affecting their stability or translation. Several recent studies have shown that miRNA regulates cell growth and apoptosis. Furthermore, clinical and experimental analyses suggested that miRNAs may function as a novel class of oncogenes or tumor suppressor genes. MicroRNA expression profiles of different tumor types, relative to their normal tissues, have recently been shown to provide phenotypic signatures for particular cancer types. Unique patterns of aberrant miRNA expression may serve as molecular biomarkers for tumor diagnosis, prognosis of disease-specific outcomes, and prediction of therapeutic responses.
Details
Supporting Data
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Figure 1. Isolation and Detection of microRNA from Urine Samples. Norgen's Urine microRNA Purification Kit was used to isolate microRNA from 8 different 1.5 mL urine samples. The purified microRNA was then used as the template in an RT-qPCR reaction to detect the human miR-21 gene. Five microlitres of the isolated RNA were used as the template in the RT step, and 5 µL from the RT step was used in the qPCR reaction. As it can be seen, the qPCR was able to successfully detect and amplify the miR-21 gene in all cases, indicating the high quality of the isolated urine microRNA. The black line in the graph above corresponds to the No Template Control.
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Kit Specifications
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Volume of Urine Processed
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0.5 - 1 mL
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| Size of RNA Purified | All sizes, including < 200 nt |
| Time to Complete 10 Purifications |
< 30 minutes
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Storage Conditions and Product Stability
All solutions should be kept tightly sealed and stored at room temperature. This kit is stable for 2 years after the date of shipment.
| Component | Cat. 29000 (25 preps) |
|---|---|
| Lysis Buffer A | 2 x 20 mL |
| Wash Solution A | 18 mL |
| Elution Solution A | 6 mL |
| Mini Spin Columns | 25 |
| Collection Tubes | 25 |
| Elution Tubes (1.7 mL) | 25 |
| Product Insert | 1 |
Documentation
FAQs
Spin Column
We recommend the use of Norgen’s Urine Preservative, which is designed for the preservation of
nucleic acids and proteins in fresh urine samples at ambient temperatures. The components of
the Urine Preservative allow samples to be stored for over 2 years at room temperature with no
detected degradation of urine DNA, RNA or proteins. Norgen’s Urine Preservative is available in
2 convenient formats: in a liquid format in Norgen’s Urine Preservative Single Dose Ampules, as
well as in a dried format in Norgen’s Urine Collection and Preservation Tubes.
A fixed speed centrifuge can be used, however reduced yields may be observed.
This may decrease the binding of the miRNA to the column.
The correct rpm can be calculated using the formula:
RPM= √RCF/(1.118x10-5)(r)
Where RCF = required gravitational acceleration (relative centrifugal force in units of g); r = radius of the rotor in cm; and RPM = the number of revolutions per minute required to achieve the necessary g-force.
RPM= √RCF/(1.118x10-5)(r)
Where RCF = required gravitational acceleration (relative centrifugal force in units of g); r = radius of the rotor in cm; and RPM = the number of revolutions per minute required to achieve the necessary g-force.
All centrifugation steps are performed at room temperature. Centrifugation at 4°C will not adversely affect kit performance.
Yes, you can. All the buffers included in this kit are in a linear relationship to the volume of urine
sample processed. Make sure that you do not deviate from the ratio specified in the product
manual. The minimum recommended urine input is 0.5 mL, and the maximum recommended
input is 1 mL. Please note that the buffers are optimized for an input of 1 mL of urine.
Adding less volume may reduce your miRNA yields. Adding more may not affect the miRNA yields EXCEPT if more Elution solution was added. Eluting miRNA in a higher volume of Elution Solution A will result in diluting your miRNA.
Your miRNA elution will be contaminated with traces of the Wash Solution A. This may dilute the
miRNA yield in the elution. Also, it may interfere with your downstream applications. Re-isolate
the eluted miRNA using the same procedure as you initially isolated the miRNA from urine but
using your elution as your input.
Some urine samples contain very little miRNA. This varies from individual to individual based on numerous factors. In order to increase the yield, the amount of urine input could be increased.
If a different Elution Solution was used other than the one provided in the kit, the buffer should be checked for any components that may interfere with the application. Common components that
are known to interfere are high salts (including EDTA), detergents and other denaturants. Check
the compatibility of your elution buffer with the intended use.
The centrifugation speed may be too low. Check the centrifuge to ensure that it is capable of generating a sufficient centrifugal force that is required to move the liquid phase through the resin. You may also spin an additional two minutes to ensure that the liquid is able to flow completely through the column.
miRNA can be degraded due to following factors:
- RNase contamination
RNases may be introduced during the use of the kit. Ensure proper procedures are followed when working with RNA. Please refer to “Working with RNA” at the beginning of this user guide.
- Procedure not performed quickly enough
In order to maintain the integrity of the RNA, it is important that the procedure be performed quickly.
- The cells are old
Older samples contain prematurely lysed cells which release RNase and can degrade RNA. Fresh urine samples are recommended.
It is recommended that the Urine samples should be centrifuged to prepare cell-free samples before freezing them.
We recommend the following steps to prepare frozen urine for isolation:
We recommend the use of Norgen’s Urine Preservative when collecting urine samples. Norgen’s Urine Preservative is designed for the preservation of nucleic acids and proteins in fresh urine samples at ambient temperatures, therefore no protein precipitation will occur and the purified nucleic acids will be of a higher quality.
We recommend the following steps to prepare frozen urine for isolation:
- Gently warm the sample to room temperature or 37°C for 5 min.
- DO NOT perform a centrifugation step after thawing frozen cell-free Urine samples - this will eliminate the precipitated proteins leading to loss of protein-bound cf-NA or exosomes.
- Proceed with the protocol.
We recommend the use of Norgen’s Urine Preservative when collecting urine samples. Norgen’s Urine Preservative is designed for the preservation of nucleic acids and proteins in fresh urine samples at ambient temperatures, therefore no protein precipitation will occur and the purified nucleic acids will be of a higher quality.
Citations
| Title | Analysis of microRNAs in Small Urinary Extracellular Vesicles and Their Potential Roles in Pathogenesis of Renal ANCA-Associated Vasculitis |
| Journal | Int. J. Mol. Sci.. 2022. |
| Authors | Jana Frydlova, Iveta Zednikova, Veronika Satrapova, Eva Pazourkova, Sarka Santorova, Zdenka Hruskova, Vladimir Tesar, Martin Vokurka, Petr Prikryl and Marie Korabecna |
| Title | Urinary Levels of miR-491-5p and miR-592 as Potential Diagnostic Biomarkers in Female Aging Patients with OAB: A Pilot Study |
| Journal | Metabolites. 2022. |
| Authors | Philippe G. Cammisotto, Abubakr H. Mossa, Samer Shamout and Lysanne Campeau |
| Title | Combined miRNA and SERS urine liquid biopsy for the point-of-care diagnosis and molecular stratification of bladder cancer |
| Journal | Molecular Medicine. 2022. |
| Authors | Tudor Moisoiu, Mihnea P. Dragomir, Stefania D. Iancu, Simon Schallenberg, Giovanni Birolo, Giulio Ferrero, Dan Burghelea, Andrei Stefancu, Ramona G. Cozan, Emilia Licarete, Alessandra Allione, Giuseppe Matullo, Gheorghita Iacob, Zoltán Bálint, Radu I. Badea, Alessio Naccarati, David Horst, Barbara Pardini, Nicolae Leopold & Florin Elec |
| Title | The 8q24 region hosts miRNAs altered in biospecimens of colorectal and bladder cancer patients |
| Journal | Cancer Medicine. 2022. |
| Authors | Amedeo Gagliardi,Giulia Francescato,Giulio Ferrero,Giovanni Birolo,Sonia Tarallo,Antonio Francavilla,Giulia Piaggeschi,Carla Di Battista,Gaetano Gallo,Alberto Realis Luc,Carlotta Sacerdote,Giuseppe Matullo,Paolo Vineis,Alessio Naccarati,Barbara Pardini |
| Title | Noninvasive Prognosis of Postmyocardial Infarction Using Urinary miRNA Ultratrace Detection Based on Single-Target DNA-Functionalized AuNPs |
| Journal | ACS Appl. Mater. Interfaces. 2018. |
| Authors | Qianlong Wang, Lancong Liu, Xiaoyi Chen, Tiantian Wang, Hua Zhou, Hui Huang, Linsen Qing, and Pei Luo |
| Title | MicroRNA isolation and quantification in cerebrospinal fluid: A comparative methodical study |
| Journal | PLOS | One. 2018. |
| Authors | Alena Kopkova, Jiri Sana, Pavel Fadrus, Tana Machackova, Marek Vecera, Vaclav Vybihal, Jaroslav Juracek, Petra Vychytilova-Faltejskova, Martin Smrcka, Ondrej Slaby |
| Title | Expression Levels of PIWI-interacting RNA, piR-823, Are Deregulated in Tumor Tissue, Blood Serum and Urine of Patients with Renal Cell Carcinoma |
| Journal | Anticancer Research. 2016. |
| Authors | Iliev R, Fedorko M, Machackova T, Mlcochova H, Svoboda M,, Pacik D, Dolezel J, Stanik M, Slaby O, |
| Title | Comparison of MicroRNA Content in Plasma and Urine Indicates the Existence of a Transrenal Passage of Selected MicroRNAs |
| Journal | Advances in Experimental Medicine and Biology. 2016. |
| Authors | Pazourkova E, Pospisilova S, Svobodova I, Horinek A, Brisuda A, Soukup V, Hrbacek J, Capoun O, Mares J, Hanus T, Babjuk M, Korabecna M. |
| Title | MicroRNAs in urine supernatant as potential non-invasive markers for bladder cancer detection |
| Journal | Neoplasma. 2016. |
| Authors | Pospisilova S, Pazourkova E, Horinek A, Brisuda A, Svobodova I, Soukup V, Hrbacek J, Capoun O, Hanus T, Mares J, Korabecna M, Babjuk M. |
| Title | Biomarker Potential of Urine miR-451 at Different Stages of Diabetic Nephropathy |
| Journal | Journal of Diabetes & Metabolism. 2016. |
| Authors | Emel Isiktas Sayilar, Mustafa Gullulu, Ercan Tuncel, Hande Peynirci, Adem Alemdar, Berrin Tunca, Unal Egeli, Gulsah Cecener, Murat Bayindir and Gokhan Cosgun |
| Title | Live Cell MicroRNA Imaging Using Exonuclease III-Aided Recycling Amplification Based on Aggregation-Induced Emission Luminogens |
| Journal | ACS Appl. Mater. Interfaces. 2016. |
| Authors | Min, X., Zhang, M., Huang, F., Lou, X., & Xia, F |
| Title | Noninvasive urinary miRNA biomarkers for early detection of pancreatic adenocarcinoma |
| Journal | American Journal of Cancer Research. 2016. |
| Authors | Debernardi S, Massat NJ, Radon T1, Sangaralingam A, Banissi A, Ennis DP, Dowe T, Chelala C, Pereira SP, Kocher HM, Young BD, Bond-Smith G, Hutchins R, Crnogorac-Jurcevic T |
| Title | MicroRNA molecular profiling from matched tumor and bio-fluids in bladder cancer |
| Journal | Molecular Cancer. 2015. |
| Authors | David A. Armstrong, Benjamin B. Green, John D. Seigne, Alan R. Schned and Carmen J. Marsit |
| Title | MicroRNAs in urine are not biomarkers of multiple myeloma |
| Journal | Journal of Negative Results in Biomedicine. 2015. |
| Authors | Lenka Sedlaříková, Lenka Bešše, Soňa Novosadová, Veronika Kubaczková, Lenka Radová, Michal Staník, Marta Krejčí, Roman Hájek1 and Sabina Ševčíková |
| Title | Feasibility of urinary microRNA detection in breast cancer patients and its potential as an innovative non-invasive biomarker. |
| Journal | BMC Cancer. 2015. |
| Authors | Thalia Erbes, Marc Hirschfeld, Gerta Rücker, Markus Jaeger, Jasmin Boas, Severine Iborra, Sebastian Mayer, Gerald Gitsch and Elmar Stickeler. |
| Title | MicroRNA-21-5p upregulation in urine samples serves as novel biomarkers for early stage renal cell carcinoma patients diagnosis |
| Journal | Cancer Res. 2014. |
| Authors | Wei Meng, Hansjuerg Alder, Kirsteen Maclean, Simon Kirste, Petra Stegmaier, Anca Grosu, Arnab Chakravarti, Tim Lautenschlaeger |
| Title | Urinary microRNA-29 correlates with urinary albumin in type 2 diabetes mellitus. |
| Journal | Chinese Journal of Nephrology. 2014. |
| Authors | Meirong Z, Hui P, Wenbo Z, Cheng W, Jun Z, Xun L, Yuanqing L, Paudel S, Qianqian W, Tanqi L. |
| Title | Urinary miR-29 Correlates with Albuminuria and Carotid Intima-Media Thickness in Type 2 Diabetes Patients. |
| Journal | PLoS One. 2013. |
| Authors | Peng H, Zhong M, Zhao W, Wang C, Zhang J, Liu X, Li Y, Paudel S, Wang Q, Lou T. |
