exosomes close up

Exosomes are small (30–150 nm) vesicles containing sophisticated RNA and protein cargo, that are constantly secreted by all cells in vitro and in vivo. Exosomes are changing research due to the intriguing functions within human body, including intercellular communication and signaling. These extracellular vesicles are growing, both in understanding their biological functions and using them towards practical applications such as development of non-invasive diagnostics and advanced therapeutics.

Dynabeads offers an array of exosome isolation products and tools for characterization and analysis of their RNA and protein content, as well as in vitro and in vivo tracing to help you further the understanding of these fascinating entities.

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Exosome testing and analysis products

Exosome isolation using Dynabeads

Ultracentrifugation protocols can be tedious and time consuming. Now it is possible to isolate intact exosomes from cell culture media or anybody fluid using the flexible and scalable Total Exosome Isolation reagents. These products and accompanying protocols are ideal for a wide range of experiments, including processing low sample volumes and handling multiple samples.

Isolation of exosome subpopulations

Total exosomes enriched from cell culture (by using the Total Exosome Isolation reagents or ultracentrifugation) can be further purified into specific subpopulations by immunomagnetic capture. Use the Dynabeads-based CD9, CD63, CD81, or EpCam-specific reagent, or combine the Streptavidin reagent with your choice of a biotinylated antibody to purify any population based on a surface antigen.

Intact exosome analysis

Analysis of exosomes is very challenging due to their small size of 30-150 nm. There are a few methods, though, which can be used for intact exosome analysis.

  • NanoSight instrument—Exosomes recovered with Total Exosome Isolation reagent can be sized and counted using a NanoSight instrument. Alternative tools are available from ParticleMetrix, Izon and Malvern.

  • Light microscopy—Exosomal RNA or membrane components can be labeled, allowing for visualization of exosomes under the microscope. Invitrogen specialized spin columns are available to remove unincorporated dye. These columns also allow for rapid buffer exchange, desalting or removal of low molecular weight (< MW 3000) contaminants.

  • Electron microscopy (EM)—One of the advantages of using Invitrogen Dynabeads magnetic separation technology to pull out pure exosomal sub-populations is that you can directly perform the EM analysis. You can analyze subsets from total exosomes enriched from cell culture using Invitrogen CD9, CD63, CD81, or EpCam-specific reagent, or use the Invitrogen Streptavidin reagent in combination with your choice of biotinylated antibody.

  • Flow cytometry—Immunomagnetic capture of exosomes on the surface of the Dynabeads allow for a clear and defined FFC/SSC detection typically in less than one hour. You can analyze subsets from total exosomes enriched from cell culture using the CD63-specific reagent, or use the Streptavidin reagent in combination with your choice of biotinylated antibody.

Exosome cargo isolation and analysis

Exosomes have been shown to transport a range of molecules between different cell types. Their cargo includes various proteins, mRNA (fragments and full length), rRNA, tRNA, miRNA, and ncRNA.

  • RNA & protein isolation—Purification of total RNA, or recovery of both protein and RNA from the same sample of pre-enriched exosomes can be achieved using the Invitrogen Total Exosome RNA and Protein Isolation Kit. The kit is compatible with all protocols for exosome isolation.

  • Protein analysis—Specific monoclonal antibodies (CD9CD63 and CD81) allow for detection of cellular and exosomal antigens by Western blot analysis. If you are looking to compare multiple protein samples on the same gel and want to keep exosomal protein complexes intact, you should check out the fast and gentle Dynabeads-based immunoprecipitation method using Invitrogen Protein A or Protein G in combination with your primary antibody of choice.

  • RNA analysis—Exosomal RNA and membrane components can be labeled using fluorescent dyes, while unincorporated dye can be easily depleted with Invitrogen spin columns. Exosomal RNA recovered using the Total Exosome RNA and Protein Isolation Kit can be analyzed by qRT-PCR using Applied Biosystems TaqMan AssaysRNA sequencing, or next-generation sequencing tools. Exosomal RNA and membrane components can be labeled using fluorescent dyes, while unincorporated dye can be removed by using Invitrogen spin columns.

Exosome-depleted Fetal Bovine Serum (FBS)

Exosome Depleted FBS

Gibco exosome-depleted FBS has the highest level of exosome depletion compared to homebrewed material and competitor FBS while retaining the vital nutrients necessary for cell culture. The work of Gibco exosome-depleted FBS has cut inefficiency that come with ultracentrifugation. Some of the product highlights include:

  • ≥90% of exosomes depleted
  • Cell culture testing of every lot
  • Specially developed for exosome research

Learn more about Gibco Exosome-Depleted FBS

KingFisher Flex

Standardization of Extra Cellular Vesicles isolation and characterization has been a highly requested need of the International Society of Extracellular Vesicles (ISEV) for several years. A highly reproducible method for pull-down and analysis was developed using the KingFisher Flex Purification System in combination with Dynabeads magnetic beads.

See how Dynabeads Exosome products are optimized on the KingFisher

Exosome isolation using Dynabeads

Ultracentrifugation protocols can be tedious and time consuming. Now it is possible to isolate intact exosomes from cell culture media or anybody fluid using the flexible and scalable Total Exosome Isolation reagents. These products and accompanying protocols are ideal for a wide range of experiments, including processing low sample volumes and handling multiple samples.

Isolation of exosome subpopulations

Total exosomes enriched from cell culture (by using the Total Exosome Isolation reagents or ultracentrifugation) can be further purified into specific subpopulations by immunomagnetic capture. Use the Dynabeads-based CD9, CD63, CD81, or EpCam-specific reagent, or combine the Streptavidin reagent with your choice of a biotinylated antibody to purify any population based on a surface antigen.

Intact exosome analysis

Analysis of exosomes is very challenging due to their small size of 30-150 nm. There are a few methods, though, which can be used for intact exosome analysis.

  • NanoSight instrument—Exosomes recovered with Total Exosome Isolation reagent can be sized and counted using a NanoSight instrument. Alternative tools are available from ParticleMetrix, Izon and Malvern.

  • Light microscopy—Exosomal RNA or membrane components can be labeled, allowing for visualization of exosomes under the microscope. Invitrogen specialized spin columns are available to remove unincorporated dye. These columns also allow for rapid buffer exchange, desalting or removal of low molecular weight (< MW 3000) contaminants.

  • Electron microscopy (EM)—One of the advantages of using Invitrogen Dynabeads magnetic separation technology to pull out pure exosomal sub-populations is that you can directly perform the EM analysis. You can analyze subsets from total exosomes enriched from cell culture using Invitrogen CD9, CD63, CD81, or EpCam-specific reagent, or use the Invitrogen Streptavidin reagent in combination with your choice of biotinylated antibody.

  • Flow cytometry—Immunomagnetic capture of exosomes on the surface of the Dynabeads allow for a clear and defined FFC/SSC detection typically in less than one hour. You can analyze subsets from total exosomes enriched from cell culture using the CD63-specific reagent, or use the Streptavidin reagent in combination with your choice of biotinylated antibody.

Exosome cargo isolation and analysis

Exosomes have been shown to transport a range of molecules between different cell types. Their cargo includes various proteins, mRNA (fragments and full length), rRNA, tRNA, miRNA, and ncRNA.

  • RNA & protein isolation—Purification of total RNA, or recovery of both protein and RNA from the same sample of pre-enriched exosomes can be achieved using the Invitrogen Total Exosome RNA and Protein Isolation Kit. The kit is compatible with all protocols for exosome isolation.

  • Protein analysis—Specific monoclonal antibodies (CD9CD63 and CD81) allow for detection of cellular and exosomal antigens by Western blot analysis. If you are looking to compare multiple protein samples on the same gel and want to keep exosomal protein complexes intact, you should check out the fast and gentle Dynabeads-based immunoprecipitation method using Invitrogen Protein A or Protein G in combination with your primary antibody of choice.

  • RNA analysis—Exosomal RNA and membrane components can be labeled using fluorescent dyes, while unincorporated dye can be easily depleted with Invitrogen spin columns. Exosomal RNA recovered using the Total Exosome RNA and Protein Isolation Kit can be analyzed by qRT-PCR using Applied Biosystems TaqMan AssaysRNA sequencing, or next-generation sequencing tools. Exosomal RNA and membrane components can be labeled using fluorescent dyes, while unincorporated dye can be removed by using Invitrogen spin columns.

Exosome-depleted Fetal Bovine Serum (FBS)

Exosome Depleted FBS

Gibco exosome-depleted FBS has the highest level of exosome depletion compared to homebrewed material and competitor FBS while retaining the vital nutrients necessary for cell culture. The work of Gibco exosome-depleted FBS has cut inefficiency that come with ultracentrifugation. Some of the product highlights include:

  • ≥90% of exosomes depleted
  • Cell culture testing of every lot
  • Specially developed for exosome research

Learn more about Gibco Exosome-Depleted FBS

KingFisher Flex

Standardization of Extra Cellular Vesicles isolation and characterization has been a highly requested need of the International Society of Extracellular Vesicles (ISEV) for several years. A highly reproducible method for pull-down and analysis was developed using the KingFisher Flex Purification System in combination with Dynabeads magnetic beads.

See how Dynabeads Exosome products are optimized on the KingFisher

Ordering information

Documentary mini-series: "Exosomes-The Next Small Thing"

We asked 10 prominent scientists to share their thoughts on exosomes research.

Video presentation: RNA Profiling of Exosomes

Presentation by Thermo Fisher Scientific scientist Sasha Vlassov. Includes a Q&A session following the presentation.

Watch video

Resources

Related videos

ISEV poster: Magnetic beads for fast and reproducible isolation/characterization of exosomes based on surface protein expression
ISEV presentation: Complete exosome workflow solution: from isolation to characterization
AGBT poster highlights: Characterization of the RNA content of the human blood-derived exosomes using the Ion Personal Genome Machine