Plasma/Serum Circulating and Exosomal RNA Purification Kits (Slurry Format)

Features and Benefits

Isolate all sizes of circulating and exosomal RNA, including microRNA
Versatile plasma and serum input volumes
Concentrate circulating and exosomal RNA into small elution volumes
Isolate inhibitor-free circulating and exosomal RNA
Bind and elute all RNA irrespective of size or GC content, without bias
Available in Spin Column or 96-well plate formats
Process 96-well plates using vacuum or centrifugation
Purified RNA is suitable for a variety of downstream applications, including Small RNA Sequencing. Find out more information on Norgen's NGS services
Compatible with Streck Cell-Free RNA BCT® Tubes
Purification is based on Norgen’s proprietary resin matrix

These kits are able to isolate all sizes of circulating and exosomal RNA, including microRNA, without the use of phenol or chloroform. The slurry format provides an advantage over other available kits in that it does not require extension tubes for the purification of free-circulating and exosomal RNA from large sample volumes. RNA can be isolated from either fresh or frozen samples using this kit. This kit is suitable for the isolation of RNA from serum or plasma prepared from blood collected only 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 including RT-PCR. The purified plasma/serum free-circulating and exosomal RNA is eluted in an elution solution that is compatible with PCR, qPCR, methylation-sensitive reverse transcription qPCR, reverse transcription PCR, Northern blotting, RNase protection and primer extension, expression array assays, and NGS.

Free-circulating plasma and serum RNA can serve as both tumor- and fetal-specific markers for cancer detection and prenatal diagnosis. As well, free-circulating RNAs have the potential to provide biomarkers for other disease states. Free-circulating RNA in plasma or serum are usually present as short fragments of less than 1000nt, and free-circulating miRNA (21nt) can also be found in plasma and serum.

Plasma/Serum Circulating and Exosomal RNA Purification Kit (Slurry)

Norgen's Plasma/Serum Circulating and Exosomal RNA Purification Kit (Slurry Format) provides a fast, reliable and simple procedure for isolating circulating RNA and exosomal RNA from plasma and serum samples ranging from 0.25 mL to 5 mL.

Plasma/Serum Circulating and Exosomal RNA Purification Mini Kit (Mini Slurry)

Norgen's Plasma/Serum Circulating and Exosomal RNA Purification Mini Kit (Slurry Format) provides a fast, reliable and simple procedure for isolating circulating and exosomal RNA from plasma and serum samples ranging from 0.25 mL to 2 mL.

Plasma/Serum Circulating and Exosomal RNA Purification 96-Well Kit (High Throughput Slurry)

Norgen's Plasma/Serum Circulating and Exosomal RNA Purification 96-Well Kit (Slurry Format) provides a high throughput method for isolating circulating and exosomal RNA from plasma and serum samples using either a vacuum manifold or centrifugation.

Plasma/Serum Circulating and Exosomal RNA Purification Maxi Kit (Maxi Slurry)

Norgen's Plasma/Serum Circulating and Exosomal RNA Purification Maxi Kit (Slurry Format) provides a fast, reliable and simple procedure for isolating circulating RNA and exosomal RNA from various amounts of plasma/serum ranging from 2 mL to 5 mL.

Details

Supporting Data

Figure 1. Isolation and Detection of Circulating RNA from Different Plasma Volumes. Norgen's Plasma/Serum Circulating RNA Purification Mini Kit (Slurry Format) was used to isolate circulating RNA from 0.5mL, 1mL and 2mL plasma. Three microlitres of the purified RNA was then used as the template in RT-qPCR reactions to detect the human 5S gene. The 5S housekeeping gene was detected from all plasma sample volumes used. The amplification of the 5S rRNA showed an increasing amount of RNA with increasing the sample input volume. This is represented by the decrease of the Ct value with increasing the sample input volume. RNA isolated from 0.5mL plasma is represented by the Red line, RNA isolated from 1mL plasma is represented by the Green line whereas RNA isolated from 2mL plasma is represented by the Blue line. The black line corresponds to the no template control.

Figure 2. Effective Isolation of Plasma Circulating RNA from Different Volumes. Norgen's Plasma/Serum Circulating RNA Purification Mini Kit (Slurry Format) was used to isolate circulating RNA from 0.5mL, 1mL and 2mL plasma. Three microlitres of the purified RNA was then used as the template in RT-qPCR reactions to detect the human 5S gene. The 5S housekeeping gene was detected from all plasma sample volumes used. The amplification of the 5S rRNA showed an increasing amount of RNA with increasing the sample input volume. This is represented by the decrease of the Ct value with increasing the sample input volume. Average Ct. values for the amplification of the 5S rRNA isolated from 0.5mL plasma is represented by the Red bar, Average Ct. values for the amplification of the 5S rRNA isolated from 1mL plasma is represented by the Green bar whereas Average Ct. values for the amplification of the 5S rRNA isolated from 2mL plasma is represented by the blue bar.

Figure 3. Effective and Consistent Detection of Plasma Exosome RNA. Norgen's Plasma/Serum Circulating and Exosomal RNA Isolation Kits can effectively isolate RNA from plasma. Plasma Exosome RNA was isolated from 500 µL of human plasma prepared from blood collected on citrate, EDTA or Heparin in triplicates using Norgen's Plasma/Serum RNA Isolation Kit (blue), a ExoQuick Exosome Precipitation Reagent (green) and Qiagen’s QIAzol extraction followed by a modified Qiagen's RNeasy Mini Kit cleanup (red). Stem loop RT-qPCR using primers specific to miR-21 and miR-16 as well as the housekeeping 5S rRNA was performed. In brief, three microliters of the 100 µL isolated RNA was then subjected to a 20 µL reverse transcription using 5S rRNA, miR-21 and miR-16 stem-loop reverse primer or reveres primer. Three microliters of the reverse transcription was used in a 20 µL real-time PCR reaction with primers to detect the human miR-21, human miR-16 and the 5S rRNA. Norgen's Plasma/Serum Exosome RNA Isolation Kit is the only product that showed consistent detection of all tested transcripts with the highest quality regardless the type of the anti-coagulant used for blood collection.

Figure 4. Isolation and Detection of Circulating RNA from Different Plasma Volumes. Norgen's Plasma/Serum Circulating RNA Purification Maxi Kit (Slurry Format) was used to isolate circulating RNA from 2mL, 3mL and 5mL plasma. Three microlitres of the purified RNA was then used as the template in RT-qPCR reactions to detect the human 5S gene. The 5S housekeeping gene was detected from all plasma sample volumes used. The amplification of the 5S rRNA showed an increasing amount of RNA with increasing the sample input volume. This is represented by the decrease of the Ct value with increasing the sample input volume. RNA isolated from 2mL plasma is represented by the Red line, RNA isolated from 3mL plasma is represented by the Green line whereas RNA isolated from 5mL plasma is represented by the Blue line. The black line corresponds to the no template control.

Figure 5. Effective Isolation of Plasma Circulating RNA from Different Volumes. Norgen's Plasma/Serum Circulating RNA Purification Maxi Kit (Slurry Format) was used to isolate circulating RNA from 2mL, 3mL and 5mL plasma. Three microlitres of the purified RNA was then used as the template in RT-qPCR reactions to detect the human 5S gene. The 5S housekeeping gene was detected from all plasma sample volumes used. The amplification of the 5S rRNA showed an increasing amount of RNA with increasing the sample input volume. This is represented by the decrease of the Ct value with increasing the sample input volume. Average Ct. values for the amplification of the 5S rRNA isolated from 2mL plasma is represented by the Red bar, Average Ct. values for the amplification of the 5S rRNA isolated from 3mL plasma is represented by the Green bar whereas Average Ct. values for the amplification of the 5S rRNA isolated from 5mL plasma is represented by the blue bar.

Kit Specifications - Spin Column
Minimum Plasma/Serum Input	0.25 mL
Maximum Plasma/Serum Input	2 mL
Size of RNA Purified	All sizes, including microRNA
Time to Complete Purification	< 40 minutes

Cat.51000 Storage Conditions and Product Stability
It is recommended to warm up Slurry C1 and Lysis Buffer A for 20 minutes at 60°C if any salt precipitation (crystallization) is observed. Slurry C1 contains grey resin that will not disappear by warming up.

Component	Cat. 51000 (50 preps)	Cat. 29500 (96 preps)	Cat. 42800 (50 preps)	Cat. 50900 (25 preps)
Slurry C1	-	-	-	6 mL
Slurry C2	12 mL	-	12 mL	-
Slurry C3	-	20 mL	-	-
Lysis Buffer A	2 x 130 mL	4 x 100 mL	2 x 130 mL	2 x 130 mL
Wash Solution A	38 mL	2 x 38 mL	38 mL	18 mL
Elution Solution A	6 mL	20 mL	6 mL	6 mL
Mini Filter Spin Columns	50	-	50	25
96-Well Filter Plate	-	1	-	-
Adhesive Tape	-	1	-	-
Collection Tubes	50	-	50	25
96-Well Collection Plate	-	1	-	-
Elution Tubes (1.7 mL)	50	-	50	25
96-Well Elution Plate	-	1	-	-
Product Insert	1	1	1	1

Documentation

RELATED BLOGS

FAQs

Mini Slurry

What If a variable speed centrifuge is not available?

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

What will happen if my centrifugation speed varied from the recommended speed?

This may lead to the degradation of the isolated RNA or reduction in the total RNA yields.

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.

Can I process a different Plasma/Serum volume?

Yes, you can. To process different Plasma/Serum volumes please check Table 1 of the protocol for the appropriate volumes that should be added from Slurry C1 (Step 1), Lysis Buffer A (Step 1) and 96-100% Ethanol (Step 3) to different Plasma/Serum sample volumes. The volume of Lysis Buffer A (Step 5) is fixed for all Plasma/Serum volumes.

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

Adding more or less from the specified volumes outlined in Table 1 of the protocol may affect both the quality and quantity of the isolated RNA.

What If I forgot to do a dry spin after my second wash?

Your elution will be contaminated with the Wash Solution A that contains Ethanol. This will dilute the RNA yield and it will interfere with your downstream applications.

Can I perform a second elution?

Yes, you can. A second elution is possible, but it is recommended that this elution is performed in a smaller volume (50 μL).

Why do my samples show low RNA yield?

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.

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

Most of the Free-Circulating Plasma/Serum RNA is degraded and present in short fragment. 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 Solution 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 solution with the intended use.

Do I need to do a DNase treatment for my RNA Elution?

You may need to do a DNase treatment to your isolated Plasma/Serum Circulating RNA. It is recommended to use Norgen’s RNase-Free DNase I Kit (Cat. 25710).

Are Norgen plasma/serum RNA purification kits compatible with blood samples from Streck Cell-Free RNA Blood Collection Tubes?

Yes, our plasma/serum RNA purification kits can be used to isolate RNA using blood samples preserved using Streck Cell-Free RNA Blood Collection Tubes.

Slurry

What If a variable speed centrifuge is not available?

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

What will happen if my centrifugation speed varied from the recommended speed?

This may lead to the degradation of the isolated RNA or reduction in the total RNA yields.

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.

Can I process a different Plasma/Serum volume?

Yes, you can. To process different Plasma/Serum volumes please check Table 1 in the protocol for the appropriate volumes of Lysis Buffer A (Step 1) and 96-100% Ethanol (Step 3) to be added to different Plasma/Serum sample volumes. The volume of Slurry C2 and Lysis Buffer A (Step 5) is fixed for all Plasma/Serum volumes.

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

Adding more or less from the specified volumes outlined in Table 1 of the protocol may affect both the quality and quantity of the isolated RNA.

What If I forgot to do a dry spin after my second wash?

Your elution will be contaminated with the Wash Solution A that contains Ethanol. This will dilute the RNA yield and it will interfere with your downstream applications.

Can I perform a second elution?

Yes, you can. A second elution is possible, but it is recommended that this elution is performed in a smaller volume (50 μL).

Why do my samples show low RNA yield?

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.

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

Most of the Free-Circulating Plasma/Serum RNA is degraded and present in short fragment. 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 Solution 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 solution with the intended use.

Do I need to do a DNase treatment for my RNA Elution?

You may need to do a DNase treatment to your isolated Plasma/Serum Circulating RNA. It is recommended to use Norgen’s RNase-Free DNase I Kit (Cat. 25710).

Are Norgen plasma/serum RNA purification kits compatible with blood samples from Streck Cell-Free RNA Blood Collection Tubes?

Yes, our plasma/serum RNA purification kits can be used to isolate RNA using blood samples preserved using Streck Cell-Free RNA Blood Collection Tubes.

High Throughput Slurry

What If a variable speed centrifuge is not available?

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

What will happen if my centrifugation speed varied from the recommended speed?

This may lead to the degradation of the isolated nucleic acids or reduction in the total nucleic acid yields.

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.

I noticed the wells are clogged. What causes this?

The wells of the 96-well plate will clog if;

There was an insufficient vacuum. Ensure that a vacuum pressure of at least -650 mbar or -25 inHg is developed
The centrifuge temperature was too low. Ensure that the centrifuge remains at room temperature throughout the procedure. Temperatures below 15°C may cause precipitates to form that can cause the wells to clog.

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

Adding more or less from the specified volumes outlined in Table 1 of the protocol may affect both the quality and quantity of the isolated RNA.

What If I forgot to do a dry spin after my second wash?

Your first elution will be contaminated with the Wash Solution. This may dilute the nucleic acid yield in your first elution and it may interfere with your down stream applications.

Can I perform a second elution?

Yes, you can. A second elution is possible, but it is recommended that this elution is performed in a smaller volume (50 μL).

Why do my samples show very low RNA yield?

Some 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.

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

If a different Elution Solution 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.

Do I need to do a DNase treatment for my RNA Elution?

You may need to do a DNase treatment to your isolated Plasma/Serum Circulating RNA. It is recommended to use Norgen’s RNase-Free DNase I Kit (Cat. 25710).

Why is my RNA degraded?

RNase contamination
RNases may be introduced during the use of the kit. Ensure proper procedures are followed when working with RNA. Please refer to “Working with RNA” at the beginning of the protocol's user guide.
Procedure not performed quickly enough
In order to maintain the integrity of the RNA, it is important that the procedure be performed quickly

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

Most of the Free-Circulating Plasma/Serum RNA is degraded and present in short fragment. The A260:280 ratio is normally between 1 – 1.6. This low A260:280 ratio will not affect any downstream application.

Is it possible to perform DNase treatment while using Slurry based High throughput Plasma/Serum Circulating and Exosomal RNA Purification Kit (Cat. 29500)?

Yes, it is possible to adapt the same supplementary protocol as provided for Cat. 42800. However, it is easier to perform this when using centrifugation based protocol for Cat. 29500 because the flowthrough can be lost at Step 4 when using vacuum manifold.

Are Norgen plasma/serum RNA purification kits compatible with blood samples from Streck Cell-Free RNA Blood Collection Tubes?

Yes, our plasma/serum RNA purification kits can be used to isolate RNA using blood samples preserved using Streck Cell-Free RNA Blood Collection Tubes.

Citations

Title	miRNA patterns in male LUSC patients - the 3-way mirror: Tissue, plasma and exosomes
Citation	Translational Oncology 2024.
Authors	Cecilia Bica a c, Ancuta Jurj a, Antonia Harangus b, Cristina Ciocan a, Alin Moldovan b, Oana Zanoaga a, Claudia Burz d e, Manuela Ferracin f, Lajos Raduly a, Ioana Berindan-Neagoe

Title	Associations of Maternal Breastmilk microRNAs and Infant Obesity Status at 1 Year
Citation	Genes 2024.
Authors	Emily Van Syoc ,Molly Stegman 3,†,Rhea Sullivan 3ORCID,Alexandra Confair 3,Kaitlyn Warren 3,4 andSteven D. Hicks

Title	Biological basis of extensive pleiotropy between blood traits and cancer risk
Citation	Genome Medicine 2024.
Authors	Miguel Angel Pardo-Cea, Xavier Farré, Anna Esteve, Joanna Palade, Roderic Espín, Francesca Mateo, Eric Alsop, Marc Alorda, Natalia Blay, Alexandra Baiges, Arzoo Shabbir, Francesc Comellas, Antonio Gómez, Montserrat Arnan, Alex Teulé, Monica Salinas, Laura Berrocal, Joan Brunet, Paula Rofes, Conxi Lázaro, Miquel Conesa, Juan Jose Rojas, Lars Velten, Wojciech Fendler, ...Miquel Angel Pujana

Title	A New Detection Method of Oral and Oropharyngeal Squamous Cell Carcinoma Based on Multivariate Analysis of Surface Enhanced Raman Spectra of Salivary Exosomes
Citation	Journal of personnalized medicine 2023.
Authors	Cosmin Ioan Faur ,Cristian Dinu, Valentin Toma ,Anca Jurj ,Radu Marginean ,Anca Onaciu ,Rare? Calin Roman ,Carina Culic, Magdalena Chirila ,Hora?iu Rotar ,Alexandra Falama?,Gabriela Fabiola ?tiufiuc ,Mihaela Hede?iu ,Oana Alma?an and Rares Ionu? ?tiufiuc

Title	Patients with abdominal aortic aneurysms have reduced levels of microRNA 122-5p in circulating exosomes
Citation	PLOS ONE 2023.
Authors	Jose L. Lopez ,Joel L. Ramirez ,Tuan Anh Phu,Phat Duong,Laura Bouchareychas,Christina R. Kuhrau,Pei-Yu Lin,Walter L. Eckalbar,Andrea J. Barczak,Joshua D. Rudolph,Lenka Maliskova,Michael S. Conte,Shant M. Vartanian,Robert L. Raffai and Adam Z. Oskowitz

Title	Mir-21 and Mir-125b as theranostic biomarkers for epithelial ovarian cancer in Tunisian women
Citation	African Health Sciences 2023.
Authors	A Habel, F Nassar, M Itani, H Bouaziz, M Hadj-Ahmed, Z Msheik, M Stayoussef, R Nasr & B Yacoubi-Loueslati

Title	MALAT1 in Liquid Biopsy: The Diagnostic and Prognostic Promise for Colorectal Cancer and Adenomas?
Citation	International Journal of General Medicine 2023.
Authors	Klara Cervena, Anna Siskova, Jiri Jungwirth, Marin Volaric, Jan Kral, Pavel Kohout, Miroslav Levy and Veronika Vymetalkova

Title	Identification of target genes and pathways related to heat tolerance in Chinese Holstein cows
Citation	Livestock Science 2023.
Authors	Rongling Li, Muhammad Jamil Ahmad, Minghai Hou, Xiao Wang, Shenhe Liu, Jianbin Li, Qiang Jiang, Jinming Huang, Liguo Yang

Title	Identification and Functional Analysis of Serum Specific miRNAs in Recurrent Aphthous Stomatitis Patients with Excess-heat or Yin-deficiency
Citation	Clinical Complementary Medicine and Pharmacology 2023.
Authors	Jie Bao a, Zhengyang Zhu b, Xizhao Zhang c, Lin Huang a, Li Xu a, Xiaobing Dou d, Yongsheng Fan a

Title	An inexpensive semi-automated sample processing pipeline for cell-free RNA extraction
Citation	nature protocols 2023.
Authors	Mira N. Moufarrej & Stephen R. Quake

Plasma/Serum Circulating and Exosomal RNA Purification Kits (Slurry Format)