Name: Urine microRNA Purification Kit
Brand: Norgen Biotek Corp.
SKU: 29000
Price: 279.000000 USD
Availability: InStock

Features and Benefits

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

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.

Kit Specifications
Volume of Urine Processed	0.5 - 1 mL
Size of RNA Purified	All sizes, including < 200 nt
Time to Complete 10 Purifications	< 30 minutes

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

RELATED BLOGS

FAQs

Spin Column

If I am not going to process my samples immediately, how should I store my samples?

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.

What if a variable speed centrifuge is not available?

A fixed speed centrifuge can be used, however reduced yields may be observed.

What will happen if my centrifugation speed varied from the recommended speed?

This may decrease the binding of the miRNA to the column.

My centrifuge speeds are defined in rpm and not in g-force. How can I convert g-force to rpm?

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.

At what temperature should I centrifuge my samples?

All centrifugation steps are performed at room temperature. Centrifugation at 4°C will not adversely affect kit performance.

Can I process a different urine volume?

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.

What if I added more or less of the specified reagents’ volume?

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.

What if I forgot to do a dry spin after my third wash?

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.

Why did my samples show very low miRNA yields?

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.

Why does my miRNA not perform well in downstream applications?

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.

What if the solutions did not flow through the column?

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.

Why is the miRNA degraded?

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.

Can I use frozen Urine samples for isolating miRNA?

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:

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	Feasibility of urinary microRNA detection in breast cancer patients and its potential as an innovative non-invasive biomarker
Citation	BMC Cancer 2015.
Authors	T Erbes, M Hirschfeld, G Rucker, M Jaeger, J Boas, S Iborra, S Mayer, G Gitsch, E Stickeler

Title	MicroRNAs in urine are not biomarkers of multiple myeloma
Citation	Journal of Negative Results in BioMedicine 2015.
Authors	Lenka Sedlaríková1,2, Lenka Bešše1,3, Sona Novosadová1 , Veronika Kubaczková1,2, Lenka Radová4 , Michal Staník5 , Marta Krejcí 6 , Roman Hájek1,2,7 and Sabina Ševcíková1,2*

Title	Noninvasive urinary miRNA biomarkers for early detection of pancreatic adenocarcinoma
Citation	Am J Cancer Res 2015.
Authors	Silvana Debernardi1 , Nathalie J Massat2 , Tomasz P Radon1 , Ajanthah Sangaralingam1 , Ana Banissi1 , Darren P Ennis1 , Thomas Dowe1 , Claude Chelala1 , Stephen P Pereira3 , Hemant M Kocher4,6, Bryan D Young5 , Giles Bond-Smith6 , Robert Hutchins6 , Tatjana Crnogorac-Jurcevic1

Title	Urinary microRNA-29 correlates with urinary albumin in type 2 diabetes mellitus
Citation	Chinese Journal of Nephrology 2014.
Authors	Z Meirong, P Hui, Z Wenbo, W Cheng, Z Jun, L Xun, L Yuanqing, SD Paudel, W Qianqian

Title	A microarray analysis of urinary microRNAs in renal disease
Citation	Clinical and Experimental Nephrology 2013.
Authors	T Konta, K Ichikawa, K Suzuki, K Kudo, H Satoh

Title	Urinary miR-29 Correlates with Albuminuria and Carotid Intima-Media Thickness in Type 2 Diabetes Patients
Citation	PLoS ONE 2013.
Authors	H Peng, M Zhong, W Zhao, C Wang, J Zhang, X Liu, Y Li, SD Paudel, Q Wang, T Lou

Urine microRNA Purification Kit