EVC Services

What We Offer

• nCS1 by Spectradyne
  • This instrument uses a technique called resistive pulse sensing (RPS) to measure your EV size distribution and concentration, which is something that needs to be calculated and reported for every EV sample you analyze so we made sure to provide a reliable and user-friendly instrument in our core.
  • RPS uses a nanopore (with sizes ranging from 80-900 nm depending on your sample) with an electric current flowing through a buffer. Your sample then runs through this pore and the amount that each EV particle disrupts this current is directly proportional to its size. It measures single EVs at a time, and if there are aggregates, they will likely clog up the pore, at which time the machine will automatically stop and allow you to reverse the flow. Your sample is also kept separate from the running buffer so there is no dilution or contamination.
  • Advantages:
  • More accurate and reproducible than more commonly used techniques such as dynamic light scattering (DLS)/nanoparticle tracking analysis (NTA). DLS/NTA is biased towards larger particles
  • DLS/NTA also fails to accurately represent polydispersions (i.e samples with heterogeneous size population), which most EV samples are after standard isolation procedures. The nCS1 will give you peaks and accurate composition reports for each size range AND the total sample averages, rather than one peak based on an average.
  • Caveats:
  • Anything lower than about 65nm is difficult to distinguish from electrical background noise.
  • Prices:
  • With technical assistance/training: $60.00 per hour ($50/hour if BYO cartridges)
  • Without technical assistance: $45.00 per hour ($35/hour if BYO cartridges)
  • User brings: sample and diluent (optional), pipettes and tips
  • Core provides: buffers and cleaning cartridge
  • User choice: Use cartridges at core or BYO cartridge
• Ultracentrifugation (UC): Extracellular Vesicle Isolation
  • UC is used for initial separation of EVs from biofluid through differential and sequential spins, including a 20,000 x g spin for larger microvesicles and concluding with a high-speed spin for those isolating “small EVs” or “exosomes” (30-150nm in size). After getting your initial EV pellet (whether high speed or low speed), UC can also be used to create a density gradient, to separate your EVs from any non-vesicular contaminating factors in your pellet.
  • Prices:
  • 20K and 100K EV pellets
  • With technical assistance/training: $40/isolation
  • Without technical assistance: $25/isolation
  • Full service: $40/isolation
    • Hourly rate is applied to sample preparation and handling in addition to centrifugation time
  • Density gradient protocol
    • With technical assistance/training: $300/sample
    • Without technical assistance: $100/sample
      
    • Full service: $300/sample
  • General Use of Centrifuges and Rotors Only
• ExoView Chip Array by NanoView Biosciences
  • For every EV preparation you make, you must characterize at least 2-3 “markers of those EVs. For small EVs the most common are the tetraspanins CD63, CD81 and CD9. The company NanoView Biosciences has created a platform called ExoView, which can simultaneously analyze EV surface expression of all three markers. The advantage is that you can put a small volume (35ul) of your biofluid (plasma, cell culure supernatant, etc) directly onto the plate for analysis without any pre-processing for EV separation. Analysis only takes one day and a half (overnight incubation, wash, and plate reading).
  • The advantage here is a quick analysis of these essential markers that are necessary to evaluate for every EV sample and biofluid you work with. This can be compared to western blotting or FACs of EV samples from the same biofluid, which is advantageous, as journals will often ask for multiple characterization techniques for publication.
  • Caveat:
  • The caveat is that you do not separate EVs based on these markers, but it gives you single EV analysis of multiple antigens. This company is also ca[able of working with individual labs to create chip arrays for antigens other than CD63, CD9 and CD81. These are also meant for human samples and have some crossover with mouse. More work is needed to evaluate crossover with other species.
  • Prices:
  • With technical assistance/training: $300/array
  • Without technical assistance: $80/array
  • Full service: $300/array
    • Prices may vary if custom capture chip is used
• Consultation
  • Consultation: study concepts, data analysis, downstream experimental design and publication/grant support
  • Part of our mission at the EV Core facility is to maximize the quality and success of publications and grant applications. We provide consultation services to provide guidance and support in experimental design, publication preparation and can also provide letters of support for grant applications.
  • Per hour: $25
• 1260 HPLC
  • 1260 Infinity II HPLC (High Performance Liquid Chromatography) by Agilent
  • The most common type of liquid chromatography for EVs is size exclusion chromatography, which uses a column-based method containing polymer beads with pores designed to trap particles of smaller sizes. Different fractions can be collected and exosomes, or small EVs, will come together in the same fraction.
  • At the core we are also prepared to explore new and innovative ways to use HPLC to separate and characterize EVs based on other parameters as your project may require.
  • Price: Price will vary depending on sample number and type/source of column used.
  • Size-exclusion HPLC: $75/hour
• BD FACSCanto by BD Biosciences: EV analysis by Flow Cytometry
  • This instrument performs flow cytometry, and can be used for EVs down to 80 nm in size. It allows for analysis of 7 different colors (surface markers) on EVs for high-throughput immune-phenotyping. This is ideal for identification of EV subsets and quantification based on fluorescence-based detection, not just light scatter. The advantage is a single-day, high throughput analysis of EVs and the ability to use your own antibodies to your molecules and species of interest.
  • Prices:
    • With technical assistance/training: $65.00/hour
    • Without technical assistance: $80.00/hour (training)
    • No technical assistance: $55.00/hour
    • Full Service: $110.00/hour
  • NOTE: For more immediate service, contact Jonni Moore at moorej@pennmedicine.upenn.edu for EV flow cytometry.
  • See https://pathbio.med.upenn.edu/pbr/portal/flowcyto/index.php for more information and pricing strategy.
• ZetaView by ParticleMetrix
  • The ZetaView instrument by ParticleMetrix can measure total EV concentration and size distribution by Nanoparticle Tracking Analysis (NTA) and also the surface charge of the particles (zeta potential). The surface charge of EVs has been linked to differences in biological behavior and is another parameter to characterize EVs for academic reviewers who require extended EV characterization for journal publications. Using the relationship between electrophoretic mobility of the particle and zeta potential, the ZetaView machine measures the zeta potential of your particles while also using NTA methods to determine EV size distribution with improved resolution and less bias towards larger particles than Nanosight. The caveat of measuring zeta potential is that it doesn’t tell you about the concentration of EVs in the sample or the EV size, but with the same ZetaView machine you can easily perform NTA and fluorescent NTA to get your concentration and size. The ZetaView machine from Particle Metrix is capable of measuring fluorescently labeled EVs, but GFP, Alexafluor-conjugated antibodies, and membrane dyes are best. FITC-conjugated antibodies photobleach too quickly for detection.
    
  • Advantages:
    •  Zeta potential, total NTA analysis and fluorescent NTA all in one machine
    • Improved resolution and less bias towards larger particles than the Nanosight
    • Less parameters for the user to manually set compared to Nanosight, increasing the amount of reproducibility between different samples and users
    • With one sample run using as low as 1-3 ul of your original EV sample, you can measure total EV size and concentration by light scatter NTA, zeta potential, and fluorescence NTA excitable by 488nm and 550nm lasers.
  • Caveats:
    • With particles below 50nm, it is still difficult to fully resolve what is background noise versus what is an actual particle, since very small EVs do not scatter light strongly. We recommend with EV samples from a new source to supplement your characterization with TEM analysis (see “integral partners” for more information on scheduling this).
  • Prices:
    • With technical assistance/training: $65.00/hour
    • Without technical assistance: $50.00/hour
    • Full Service: $65.00/hour