Cell Culture Media Exosome Purification and RNA Isolation Kits

Features and Benefits

Purification and enrichment of intact cell culture media exosomes for functional studies
Isolate all sizes of RNA, including microRNA, irrespective of size or GC content, without bias.
Versatile cell culture media input volumes
No phenol extractions, Proteinase K treatment, nor carrier RNA required
No time-consuming ultracentrifugation, filtration nor special syringes required
No precipitation reagents nor overnight incubation required
Pure exosomes are purified and are free from any other RNA-binding proteins
Purification is based on spin column chromatography that uses Norgen’s proprietary resin
The purified RNA is suitable for a variety of downstream applications, including Small RNA Sequencing. Find out more information on Norgen's NGS services

Norgen's Cell Culture Media Exosome Purification and RNA Isolation Kits constitute an all-in-one system for the purification of exosomes and the subsequent isolation of exosomal RNA from different cell culture media sample volumes. The purification is based on spin column chromatography that employs Norgen’s proprietary resin. The kit is 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 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. 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.

Cell Culture Media Exosome Purification and RNA Isolation Mini Kit

For sample input volumes ranging from 5 mL to 10 mL.

Cell Culture Media Exosome Purification and RNA Isolation Midi Kit

For sample input volumes ranging from 10 mL to 20 mL.

Cell Culture Media Exosome Purification and RNA Isolation Maxi Kit

For sample input volumes ranging from 20 mL to 35 mL.

Details

Supporting Data

Figure 1. Isolation of RNA from exosomes purified from different cell culture media volumes. Norgen's Cell Culture Media Exosome Purification and RNA Isolation Mini Kit (Cat# 60700) was used to isolate exosomal RNA from exosomes purified from different cell culture media 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 exosomal RNA (A) The avg. Ct value for exosomal miR-26a is linearly decreasing with increasing the sample input volume. (B) The relative amount of the exosomal miR-26a shows excellent linearity with a percentage of recovery of more than 90%

Figure 2. Isolation of RNA from exosomes purified from different cell culture media volumes. Norgen's Cell Culture Media Exosome Purification and RNA Isolation Midi Kit (Cat# 60800) was used to isolate exosomal RNA from exosomes purified from different cell culture media 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 isolatedexosomal RNA. (A) The avg. Ct value for exosomal miR-26a is linearly decreasing with increasing the sample input volume. (B) The relative amount of the exosomal miR-26a shows excellent linearity with a percentage of recovery of more than 90%.

Figure 3. Isolation of RNA from exosomes purified from different cell culture media volumes. Norgen's Cell Culture Media Exosome Purification and RNA Isolation Maxi Kit (Cat# 60900) was used to isolate exosomal RNA from different cell culture media volumes from exosomes purified 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 exosomal RNA. (A) The avg. Ct value for exosomal miR-26a is linearly decreasing with increasing the sample input volume. (B) The relative amount of the exosomal miR-26a shows excellent linearity with a percentage of recovery of more than 90%.

Kit Specifications
Sample Type	Cell-Free Cell Culture Media
Sample Volume Range	5 mL to 10 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 to 40 minutes
Average Yields*	Variable depending on specimen

* Please check page 4 of the product insert for Average Yields and 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 from the date of shipment. It is recommended to warm Lysis Buffer A for 20 minutes at 60°C if any salt precipitation is observed.

Component	Cat. 60700 (50 preps)	Cat. 60800 (25 preps)	Cat. 60900 (15 preps)
Slurry E	12.5 mL	12.5 mL	12.5 mL
ExoC Buffer	1.5 mL	1.5 mL	1.5 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 Column	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

Documentation

RELATED BLOGS

FAQs

Mini, Midi, Maxi

What If a variable speed centrifuge is not available?

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

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.

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

Adding more or less than the specified volumes may reduce both the quality and the quantity of the purified RNA. Eluting your RNA in high volumes will increase the yield but will lower the concentration. Eluting in small volumes will increase the concentration but will lower the overall yield.

What if I forgot to do a dry spin before my final elution step?

Your purified RNA will be contaminated with the Wash Solution A. This may reduce the quality of your purified RNA and will interfere with your downstream applications.

Can I perform a second elution?

Yes, but it is recommended that the 2nd elution be in a smaller volume (50% of 1st Elution). It is also recommended to perform the 2nd elution into a separate elution tube to avoid diluting the 1st elution.

Why do my samples show low RNA yield?

Exosomes contain very little RNA. This varies significantly depending on the number of exosomes that are shed from the cultured cells, as well as the kind of treatment the cultured cells are receiving. In order to increase the yield, the amount of media could be increased.

Why is the A260/280 ratio of the purified RNA lower than 2.0?

Most of the Exosomal RNA is short RNA fragments with a very low concentration where the A260/280 ratio tends to decrease with the decrease in the RNA concentration. The A260/280 ratio is normally between 1-1.6. This low A260/280 ratio will not affect any downstream application.

Why does my isolated RNA not perform well in downstream applications?

If a different Elution Buffer 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.

Citations

Title	Adipose-derived stem cell exosomes act as delivery vehicles of microRNAs in a dog model of chronic hepatitis
Citation	Nanotheranostics 2023.
Authors	Ilaria Zanolla1, Martina Trentini2, Elena Tiengo2, Federica Zanotti2, Tommaso Pusceddu2, Andrea Rubini3, Giuseppe Rubini4, Francesca Brugnoli4, Danilo Licastro5, Marco Debortoli5, Lucia Gemma Delogu6, Letizia Ferroni6, Luca Lovatti2, Barbara Zavan

Title	Effects of microgravity on neural crest stem cells
Citation	Frontiers in Neuroscience 2023.
Authors	Yilin Han1† , Povilas Barasa2† , Lukas Zeger1†, Sara B. Salomonsson3‡ , Federica Zanotti4‡ , Marcel Egli5,6, Barbara Zavan4 , Martina Trentini4 , Gunnar Florin7 , Alf Vaerneus7, Håkan Aldskogius1 , Robert Fredriksson3 and Elena N. Kozlova1*

Title	RNA-Binding Protein Motifs Predict microRNA Secretion and Cellular Retention in Hypothalamic and Other Cell Types
Citation	Biomedicnes 2023.
Authors	Wenyuan He 1 and Denise D. Belsham

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Cell Culture Media Exosome Purification and RNA Isolation Kits