Research Applications

Nanomembranes for Extracellular Vesicle Visualization

Capture, image, and analyze tagged single extracellular vesicles using our innovative nanomembrane-based technology, opening the door to exciting Research Use Only and Invitro diagnostics.

The EV Visualization Concept. A) 3D rendering showing the one-by-one capture of EVs in the pores of a SiMPore nanomembrane, with fluorescent affinity tagged EVs appearing in bright green. B) The Microfluidic device enabled by Silicon Membranes for EV Visualization (the µSiM-EV).

What are Extracellular Vesicles?

Extracellular vesicles (EVs) are lipid bilayer-bound vesicles that emerge from nearly all cell types of the mammalian body. EVs carry cargo from their cells of origin, thereby making them candidate markers for cancer and other diseases.

EVs also play significant roles in cell-to-cell communication in diverse tissues. These vesicles are heterogeneous in nature. Even EVs originating from the same cell often have different characteristics (e.g. size, shape, biomolecular composition), emphasizing the need and importance of single EV analyses over bulk analyses of EV populations, which would mask their inherent heterogeneity.

Types of Extracellular Vesicles:

According to MISEV 2018 criteria, the two EV classes of greatest interest fall into two broad categories based on size and biological origin:

• Small EVs (sEVs; a.k.a., exosomes) that are 30–200 nm in diameter and thought to originate from the endosomal pathway
• Medium and large EVs (a.k.a., microvesicles, ectosomes) that are 200–1,000 nm in diameter and thought to originate from budding of the plasma membrane

Single Extracellular Vesicle Analysis Made Easy

Count 1,000s individual, marker-positive EVs using conventional microscopy

µSiM-EV Options of Our Microfluidics enabled by Silicon Membranes for Extracellular Vesicle Visualization

Our innovative nanomembrane technology allows imaging of antibody-tagged and/or dye-labeled EVs by optical microscopy using conventional epifluorescent inverted and/or confocal microscopy. No dedicated and expensive instrumentation is needed. Simply add your prepared EV sample and image.

The µSiM-EV works with any sample with only minimal centrifugation to remove cellular debris or with more involved samples prepared by size-exclusion chromatography and/or ultracentrifugation. Also compatible with common immunoassay and other typical buffers.

Easy to Use

Simple hand-pipette sample loading is made easy by the µSiM-EV Loading Accessory.

Sample Loading and Imaging. A) Samples are loaded by manual pipette into the µSiM-EV as shown. Fluid injection forces the sample bearing EVs up into the center well, thereby capturing EVs on the well’s integrated nanomembrane. B) The µSiM-EV is then transferred to a holder for imaging on a conventional epifluorescent microscope.

Single EV Image-based Analyses

Optically transparent nanomembranes with EV-sized pores efficiently capture 100,000s of EVs, enabling robust digital EV analyses with statistical confidence.

Immunolabeling Analysis by µSiM-EV. µSiM-EV analysis of EVs prepared by Size-Exclusion Chromatography. 0.5 mL human plasma applied to a column comprising 10 mL of Sepharose CL-4B and 2 mL FractoGel-sulfate A) CFSE (cyan) and anti-CD63 (magenta) fluorescent micrographs are shown from representative fields-of-view from across the µSiM-EV’s membrane. B) Particle counts are rendered from ImageJ Comdet. 5pxl max separation 3pxl/450nm max particle size, 3 intensity threshold.

Dual-Mode Imaging

Prepare samples and then image by scanning electron microscopy (SEM) on the same devices and correlate optical and electron micrographs.

Optical and Electron Microscopy Imaging with the µSiM-EV. Representative fluorescent and scanning electron micrographs are shown for 850 nm diameter particles captured with the pores of a 500 nm cut-off nanomembrane (left set) and for 50 nm diameter particles captured in the pores of a 50 nm cut-off nanomembrane (right set).

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