EXTRAClean Plasma/Serum RNA Purification Kit
EXTRAClean Plasma/Serum RNA Purification Kit
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Overview
- Versatile plasma/serum input ranges
- No phenol extractions
- Fast and easy processing of samples for the purification and the isolation of RNA.
- No time-consuming ultracentrifugation.
- More effective than any other method.
- EXTRAClean resin separation matrix provides highly purified RNA
These kits provide a fast, reliable and convenient method to purify and concentrate high quality, high purity and inhibitor-free cell-free circulating and exosomal RNA using a convenient spin column method. These kits can purify RNA from fresh or frozen serum or plasma samples prepared from blood collected on either EDTA or Citrate. Plasma samples prepared from blood collected on heparin should not be used, as heparin can significantly interfere with many downstream applications such as RT-PCR. The purified plasma/serum RNA is fully compatible with all downstream applications including PCR, qPCR, methylation-sensitive reverse transcription qPCR, reverse transcription PCR, Northern blotting, RNase protection and primer extension, expression array assays, and NGS. The EXTRAClean columns undergo stringent processing and rigorous quality control measures to minimize contamination traces, ensuring optimal results for sensitive applications such as NGS.
Background
Cell-free circulating RNA, including exosomal RNA in plasma or serum, has the potential to provide biomarkers for certain cancers and disease states, and includes tumor-specific extracellular RNA in the blood. Exosomes are 40 – 100 nm membrane vesicles, which are secreted by most cell types. Exosomes can be found in saliva, blood, urine, amniotic fluid and malignant ascitic fluids, among other biological fluids. Evidence has been accumulating recently that these vesicles act as cellular messengers, conveying information to distant cells and tissues within the body. The exosomes contain cell-specific proteins, lipids and RNAs, which are transported to other cells, where they can alter function and/or physiology. These exosomes may play a functional role in mediating adaptive immune responses to infectious agents and tumours, tissue repair, neural communication and transfer of pathogenic proteins. Recent work has demonstrated the presence of distinct subsets of microRNAs within exosomes which depend upon the tumour cell type from which they are secreted. For this reason, exosomal RNAs may serve as biomarkers for various diseases including cancer. As the RNA molecules encapsulated within exosomes are protected from degradation by RNAses, they can be efficiently recovered from biological fluids, such as plasma or serum.
EXTRAClean Plasma/Serum RNA Purification Mini Kit
This kit can purify RNA from fresh or frozen serum or plasma samples prepared from blood collected on either EDTA or Citrate, from volumes ranging from 50 μL to 200 μL. The purified plasma/serum RNA is eluted in a flexible final volume of 10 μL to 25 μL.
EXTRAClean Plasma/Serum RNA Purification Midi Kit
This utilizes a two-column method, and can purify RNA from fresh or frozen serum or plasma samples prepared from blood collected on either EDTA or Citrate, from volumes ranging from 250 μL to 1.5 mL. The first column will handle the large volume input of bodily fluids that is followed by a concentration on a mini column for a final elution of 50 μL to 100 μL.
EXTRAClean Plasma/Serum RNA Purification Maxi Kit
This kit can purify RNA from fresh or frozen serum or plasma samples prepared from blood collected on either EDTA or Citrate, from volumes ranging from 2 mL to 5 mL. The first column will handle the large volume input of bodily fluids that is followed by a concentration on a mini column for a final elution of 50 μL to 100 μL.
Details
Supporting Data
Kit Specifications | |
Sample Type | Plasma/Serum |
Anti-Coagulant (for Plasma)† | EDTA or Citrate |
Sample Volume Range | 2 to 5 mL |
Minimum Elution Volume | 50 μL |
Maximum Elution Volume | 100 μL |
Time to Complete 10 Purifications | 35 – 40 minutes |
Size of RNA Purified | All sizes, including miRNA and small RNA (<200 nt) |
Average Yields¥ | Variable depending on specimen |
† This kit is suitable for the isolation of RNA from fresh or frozen serum or plasma prepared from blood collected on either EDTA or Citrate. Plasma samples prepared from blood collected on heparin should not be used as heparin can significantly interfere with many downstream applications such as RT-PCR
¥ Please check page 5 for Average Plasma/Serum Yields and Common RNA Quantification Methods
Storage Conditions and Product Stability
All solutions 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.
Component | Cat. 73700 (50 preps) | Cat. 73710 (20 preps) | Cat. 73720 (10 preps) |
---|---|---|---|
Lysis Buffer A | 2 x 20 mL | 100 mL | 1 x 130 mL 1 x 30 mL |
Wash Solution A | 18 mL* | 1 x 38 mL* 1 x 18 mL* |
38 mL* |
Elution Solution A | 6 mL | 6 mL | 6 mL |
Elution Buffer F | - | - | 15 mL |
EXTRACLean Micro Spin Columns | 50 | - | - |
EXTRACLean Mini Spin Columns | - | 20 | 10 |
EXTRACLean Midi Spin Columns | - | 20 | - |
EXTRACLean Maxi Spin Columns | - | - | 10 |
Collection Tubes | - | 20 | 10 |
Elution Tubes (1.7 mL) | 50 | 20 | 10 |
Product Insert | 1 | 1 | 1 |
* For the preparation of working solutions, please see important Notes (Notes prior to use)
Documentation
FAQs
Mini, Midi, Maxi
A fixed speed centrifuge can be used, however reduced yields may be observed.
All centrifugation steps are performed at room temperature. Centrifugation at 4°C will not adversely affect kit performance.
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.
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
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
Plasma/Serum samples contain very little RNA. This varies from individual to individual based on numerous variables. In order to increase the yield, the amount of Plasma/Serum input could be increased.
Most of the Free-Circulating Plasma/Serum RNA is short RNA fragments. The A260:280 ratio is normally between 1 – 1.6. This low A260:280 ratio will not affect any downstream application.
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.
You may need to do a DNase treatment to your isolated Plasma/Serum miRNA. It is recommended to use Norgen’s RNase-Free DNase I Kit (Cat# 25710). Also please refer to the protocol for optional on-column DNA removal outlined in the protocols.