All-in-one system for the purification of exosomes and the subsequent isolation of RNA from urine samples
For research use only and NOT intended for in vitro diagnostics.
All-in-one system for the purification of exosomes and the subsequent isolation of RNA from urine samples
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Norgen’s Exosome Purification and RNA Isolation Kits constitute all-in-one systems for the purification of exosomes and the subsequent isolation of exosomal 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 in that they do not require any special instrumentation, protein precipitation reagents, extension tubes, phenol/chloroform or protease treatments. Moreover, the kit allows the user to elute into a flexible elution volume ranging from 50 µL to 100 µL. The purified RNA is free from any protein-bound circulating RNA and of the highest integrity. 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 Purification and RNA Isolation Mini Kit
For sample volumes ranging from 250 µL to 1 mL.
Urine Exosome Purification and RNA Isolation Midi Kit
For sample volumes ranging from 2 mL to 10 mL.
Urine Exosome Purification and RNA Isolation Maxi Kit
For sample volumes ranging from 11 mL to 30 mL.
Figure 1. Isolation of RNA from exosomes purified from different urine volumes. Norgen's Urine Exosome Purification and RNA Isolation Mini Kit (Cat# 58400) was used to isolate RNA from exosomes purified from different urine volumes using the same kit. 2 µL 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 avg. Ct value for 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. Comparing exosome yields from different commercial purification methods. Intact exosomes were purified from 5 mL urine using Norgen's Urine Exosome Purification and RNA Isolation Midi Kit (Cat# 58700), Competitor S's kit, and Competitor L's kit. Exosomes purified using Norgen's kit were resuspended in 400 µL Norgen's ExoR buffer, diluted 1:1,000 and visualized on the NanoSight LM10 instrument. The analysis shows that Norgens kit isolated 115 nm exosomes with a recovery of 8.36 x 109 particles/mL urine samples. No impurities were found to be contaminating the exosomes purified using Norgen's Urine Exosome Purification and RNA Isolation Midi kit. Additionally, exosomes with a broader size range covering from 75nm to 250nm were purified from 5 mL urine with a higher concentration as compared to the other two methods.
Figure 3. Comparing exosome profiles from Norgen’s method and ultracentrifugation. Exosomes purified using Norgens kit and ultracentrifugation were resuspended in 400 µL of Norgens ExoR buffer, diluted 1:1,000 and visualized on the NanoSight LM10 instrument. The analysis shows that Norgens kit purified exosomes with sizes ranging from 65 nm - 195 nm, with a total recovery of 7.63 x 108 particles/mL. No impurities were found to be contaminating the exosomes purified using Norgen's Urine Exosome Purification and RNA Isolation Midi Kit as opposed to the exosomes purified using ultracentrifugation, which purified exosomes with larger particle sizes ranging from 125 nm to 235 nm with a total recovery of 1.56 x 108 particles/mL.
Figure 4. Isolation of RNA from exosomes purified from different urine volumes. Norgen's Urine Exosome Purification and RNA Isolation Midi Kit (Cat# 58700) was used to isolate RNA from exosomes purified from different urine volumes using the same kit. 2 µL 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 avg. Ct value for 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 5. Isolation of RNA from exosomes purified from different urine volumes. Norgen's Urine Exosome Purification and RNA Isolation Maxi Kit (Cat# 58800) was used to isolate RNA from exosomes purified from different urine volumes using the same kit. 2 µL 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 avg. Ct value for 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%.
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Kit Specifications
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| Minimum Urine Input |
250 μL
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Maximum Urine Input
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1 mL
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Size of Exosomes Purified
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40 nm - 150 nm
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| 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
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| 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. 58400 (50 preps) | Cat. 58700 (25 preps) | Cat. 58800 (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 | 20 mL | 20 mL | 20 mL |
| Lysis Additive B | 2 mL | 2 mL | 2 mL |
| Wash Solution A | 18 mL | 18 mL | 18 mL |
| Elution Solution A | 6 mL | 6 mL | 6 mL |
| Mini Filter Spin Columns | 50 | 25 | 15 |
| Mini Spin Columns | 50 | 25 | 15 |
| Collection Tubes | 50 | 25 | 15 |
| Elution Tubes (1.7 mL) | 100 | 50 | 30 |
| Product Insert | 1 | 1 | 1 |
Midi (58700)
| Title | Placental exosomes isolated from urine of patients with gestational diabetes exhibit a differential profile expression of microRNAs across gestation |
| Journal | International Journal of Molecular Medicine. 2020. |
| Authors | Ana Sofía Herrera-Van Oostdam Juan Carlos Toro-Ortíz, et al. |
| Title | Comparing small urinary extracellular vesicle purification methods with a view to RNA sequencing—Enabling robust and non-invasive biomarker research |
| Journal | Biomolecular Detection and Quantification. 2019. |
| Authors | Veronika Mussack, Georg Wittmann, Michael W. Pfaffl |
Maxi (58800)
| Title | A Pilot Study of miRNA Expression Profile as a Liquid Biopsy for Full-Marathon Participants |
| Journal | Sports. 2021. |
| Authors | Tomoaki Kuji, Takehito Sugasawa, Shin-ichiro Fujita, Seiko Ono, Yasushi Kawakami and Kazuhiro Takekoshi |
