Urine Exosome and Free-Circulating RNA Isolation Kits

Features and Benefits

Isolate all sizes of RNA, including microRNA, irrespective of size or GC content, without bias
Versatile sample input ranges
Isolate all sizes of free-circulating RNA, including microRNA
Bind and elute all RNA irrespective of size or GC content, without bias
The purified exosomal RNA is free from any circulating RNA-binding proteins
No phenol extractions, Proteinase K treatment nor carrier RNA required
No time-consuming ultracentrifugation, filtration nor special syringes are required
No precipitation reagents nor overnight incubation required
Concentrate isolated exosomal RNA and free-circulating RNA into a flexible elution volume ranging from 50 µL to 100 µL
Purification is based on spin column chromatography that uses Norgen’s proprietary resin separation matrix

These kits provide a fast, reliable and convenient method to sequentially isolate and concentrate exosomal RNA as well as Free-Circulating RNA from different urine sample volumes. The purification is based on spin column chromatography that employs Norgen’s proprietary resin. These kits are designed to isolate all sizes of RNA, including microRNA, irrespective of size or GC content, without bias. These kits provide a clear advantage over other available kits since they do not require any special instrumentation, protein precipitation reagents, extension tubes, phenol/chloroform or protease treatments. Moreover, the kits allow the user to elute into a flexible elution volume ranging from 50 µL to 100 µL. The RNA isolated from the purified exosomes is free from any protein-bound circulating RNA and is of the highest integrity. Moreover, the free-circulating, protein-bound, RNA is free from any exosomal RNA. The purified RNA 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.

Urine Exosome and Free-Circulating RNA Isolation Mini Kit

For sample volumes ranging from 250 µL to 1 mL.

Urine Exosome and Free-Circulating RNA Isolation Midi Kit

For sample volumes ranging from 2 mL to 10 mL.

Urine Exosome and Free-Circulating RNA Isolation Maxi Kit

For sample volumes ranging from 11 mL to 30 mL.

Details

Supporting Data

Figure 1. Isolation of RNA from exosomes purified from different urine volumes. Norgen’s Urine Exosome and Free-Circulating RNA Isolation Mini Kit (Cat# 59200) was used to isolate exosomal RNA from different urine volumes ranging from 0.25 mL and up to 1 mL. Two microlitres of the isolated RNA was then used as the template in RT-qPCR reactions to assess the amplification of the isolated urinary exosomal miR-30a. (A) The urinary exosomal miR-30a is linearly decreasing with increasing the sample input volume. B) The relative amount of the urinary exosomal miR-30a shows excellent linearity with a percentage of recovery of more than 90%.

Figure 2. Isolation of Free-Circulating RNA after the isolation of Exosomal RNA from different urine volumes. Norgen’s Urine Exosome and Free-Circulating RNA Isolation Mini Kit (Cat# 59200) was used to isolate Free-Circulating RNA after the isolation of exosomal RNA from different urine volumes ranging from 0.25 mL and up to 1 mL. Two microlitres of the isolated RNA was then used as the template in RT-qPCR reactions to assess the amplification of the isolated urinary free-circulating miR-21. (A) The free-circulating miR-21 is linearly decreasing with increasing the sample input volume. B) The relative amount of the free-circulating miR-21 shows excellent linearity with a percentage of recovery of more than 90%.

Figure 3. Isolation of RNA from exosomes purified from different urine volumes. Norgen’s Urine Exosome and Free-Circulating RNA Isolation Midi Kit (Cat# 59300) was used to isolate Exosomal RNA from different urine volumes ranging from 2 mL and up to 10 mL. Two microlitres of the isolated RNA was then used as the template in RT-qPCR reactions to assess the amplification of the isolated urinary exosomal miR-30a. (A) The urinary exosomal miR-30a is linearly decreasing with increasing the sample input volume. B) The relative amount of the urinary exosomal miR-30a shows excellent linearity with a percentage of recovery of more than 90%.

Figure 4. Isolation of Free-Circulating RNA after the isolation of Exosomal RNA from different urine volumes. Norgen’s Urine Exosome and Free-Circulating RNA Isolation Midi Kit (Cat# 59300) was used to isolate Free-Circulating RNA from after the isolation of Exosomal RNA from different urine volumes ranging from 2 mL and up to 10 mL. Two microlitres of the isolated RNA was then used as the template in RT-qPCR reactions to assess the amplification of the isolated urinary free-circulating miR-21. (A) The free-circulating miR-21 is linearly decreasing with increasing the sample input volume. B) The relative amount of the free-circulating miR-21 shows excellent linearity with a percentage of recovery of more than 90%.

Figure 5. Isolation of RNA from exosomes purified from different urine volumes. Norgen’s Urine Exosome and Free-Circulating RNA Isolation Maxi Kit (Cat# 59400) was used to isolate exosomal RNA from different urine volumes ranging from 11 mL to 30 mL. Two microlitres of the isolated RNA was then used as the template in RT-qPCR reactions to assess the amplification of the isolated urinary exosomal miR-30a. (A) The urinary exosomal miR-30a is linearly decreasing with increasing the sample input volume. B) The relative amount of the urinary exosomal miR-30a shows excellent linearity with a percentage of recovery of more than 90%.

Figure 6. Isolation of Free-Circulating RNA after the isolation of Exosomal RNA from different urine volumes. Norgen’s Urine Exosome and Free-Circulating RNA Isolation Maxi Kit (Cat# 59400) was used to isolate Free-Circulating RNA after the isolation of exosomal RNA from different urine volumes ranging from 11 mL to 30 mL. Two microlitres of the isolated RNA was then used as the template in RT-qPCR reactions to assess the amplification of the isolated urinary free-circulating miR-21. (A) The free-circulating miR-21 is linearly decreasing with increasing the sample input volume. B) The relative amount of the free-circulating miR-21 shows excellent linearity with a percentage of recovery of more than 90%.

Kit Specifications
Minimum Urine Input	250 µL
Maximum Urine Input	1 mL
Size of RNA Purified	All sizes, including miRNA and small RNA (< 200 nt)
Elution Volume	50 - 100 µL
Time to Complete 10 Purifications	35 - 40 minutes
Average Yields*	Variable depending on specimen

*Please check page 4 of the product insert for the average yields and the common RNA quantification methods.

Storage Conditions and Product Stability
All buffers should be kept tightly sealed and stored at room temperature. This kit is stable for 2 years after the date of shipment. It is recommended to warm Lysis Buffer A for 20 minutes at 60°C if any salt precipitation is observed.

Important Note
Urine samples stored at -70°C, -20°C or at 4°C will develop some precipitation due to the aggregation of some of the highly abundant proteins in urine. Eliminating these precipitates using centrifugation or filtration may cause the loss of exosomes. Furthermore, these precipitates may affect the quality of the purified nucleic acid. We recommend the use of Norgen’s Urine Preservative when collecting urine samples, 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 as 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.

Component	Cat. 59200 (50 preps)	Cat. 59300 (25 preps)	Cat. 59400 (15 preps)
Slurry E	12.5 mL	12.5 mL	12.5 mL
ExoC Buffer	8 mL	30 mL	50 mL
ExoR Buffer	12 mL	12 mL	12 mL
Lysis Buffer A	2 x 20 mL	2 x 20 mL	2 x 20 mL
Lysis Additive B	2 mL	2 mL	2 mL
Wash Solution A	2 x 18 mL	18 mL	18 mL
Elution Solution A	2 x 6 mL	6 mL	6 mL
Mini Filter Spin Columns	50	25	15
Mini Spin Columns	100	50	30
Collection Tubes	100	50	30
Elution Tubes (1.7 mL)	100	50	30
Product Insert	1	1	1

Documentation

PROTOCOLS

(59200) Urine Exosome and Free-Circulating RNA Isolation Mini Kit - Protocol (50 prep)

(59300) Urine Exosome and Free-Circulating RNA Isolation Midi Kit - Protocol (25 prep)

(59400) Urine Exosome and Free-Circulating RNA Isolation Maxi Kit - Protocol (15 prep)

Version Available Here.

Urine Exosome and Free-Circulating RNA Isolation Kits