Cell Culture Media Exosome Purification and RNA Isolation Kits

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.

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Cell Culture Media Exosome Purification and RNA Isolation Kits