Urine Exfoliated Cell RNA Purification Kit

Supplier
Norgen Biotek
Cat. no.
1022500
Kit size
20
Links
Norgen Biotek Protocol Information sheet

Features

• Fast processing -Rapid spin-column format allows for the processing of multiple samples in 20 minutes.
• Isolate a diversity of RNA species - All RNA species can be isolated, from large mRNA and ribosomal RNA down to microRNA and small interfering RNA.
• Isolate RNA from small samples - RNA can be isolated and detected from as little as 100 exfoliated cells.
• No phenol:chloroform extractions - Total exfoliated cell RNA is isolated without the use of harmful chemicals such as phenol or chloroform.
• Recovered RNA is suitable for downstream applications - Purified RNA is fully compatible with real time PCR, reverse transcription PCR, Northern blotting, RNAse protection and primer extension, and expression array assays.

Urine (Exfoliated Cell) RNA Purification Kit provides a rapid method for the isolation and purification of total RNA from exfoliated cells that have been shed into the urine from the urinary tract. RNA biomarkers from exfoliated cells can be used as non-invasive tool for a number of diagnostic and research applications including the diagnosis and monitoring of bladder, kidney, or other urinary-tract cancers. The kit purifies all sizes of RNA, from large mRNA and ribosomal RNA down to microRNA (miRNA) and small interfering RNA (siRNA). The RNA is preferentially purified from other cellular components such as proteins, as well as from the contaminating species found in urine such as glucose and salts, without the use of phenol or chloroform. The purified RNA is of the highest integrity, and can be used in a number of downstream applications including real time PCR, reverse transcription PCR, Northern blotting, RNase protection and primer extension, and expression array assays.

Purification is based on spin column chromatography using Norgen’s proprietary resin as the separation matrix. The RNA is preferentially purified from other cellular components such as proteins without the use of phenol or chloroform. The process involves first lysing the exfoliated cells with the provided Lysis Solution (please see the flow chart on page 4). Ethanol is then added to the lysate, and the solution is loaded onto a spin-column. Norgen’s resin binds RNA in a manner that depends on ionic concentrations, thus only the RNA will bind to the column while the contaminating proteins will be removed in the flowthrough or retained on the top of the resin. The bound RNA is then washed twice with the provided wash buffer in order to remove any remaining impurities, and the purified total RNA is eluted with the elution buffer. The purified RNA is of the highest integrity, and can be used in a number of downstream applications.

Column Binding Capacity	50 µg
Volume of Urine Processed	1 – 50 ml
Maximum Input of Exfoliated Cells	1 x 10exp6
Size of RNA Purified	All sizes, including small RNA (<200 nt)
Time to Complete 10 Purifications	20 minutes
Average Yields	~ 1 µg RNA per 1x10exp5cells	(Varies due to cell density sample)

Isolation and Detection of RNA from the Exfoliated Cells in 50 mL of Urine

RNA was isolated from the exfoliated cells in a 50 mL urine sample obtained from a healthy male using Norgen’s Urine (Exfoliated Cell) RNA Purification Kit. The isolated RNA was then subjected to reverse transcription and real-time PCR using primers to detect the ribosomal S14 gene. The red lines in the PCR baseline graph above correspond to RNA standards, while the blue lines correspond to the successful PCR results when RNA isolated from the exfoliated cells in 50 mL of urine were used as the template.

User-Friendly Procedure

• Quantitative, real-time RT-PCR
• RT-PCR
• Northern blotting
• RNase protection and primer extension
• Expression array assays