A Label-Free Electrical Impedance Spectroscopy for Detection of Clusters of Extracellular Vesicles Based on Their Unique Dielectric Properties

Abstract

Extracellular vesicles (EVs) have gained considerable attention as vital circulating biomarkers since their structure and composition resemble the originating cells. The investigation of EVs’ biochemical and biophysical properties is of great importance to map them to their parental cells and to better understand their functionalities. In this study, a novel frequency-dependent impedance measurement system has been developed to characterize EVs based on their unique dielectric properties. The system is composed of an insulator-based dielectrophoretic (iDEP) device to entrap and immobilize a cluster of vesicles followed by utilizing electrical impedance spectroscopy (EIS) to measure their impedance at a wide frequency spectrum, aiming to analyze both their membrane and cytosolic charge-dependent contents. The EIS was initially utilized to detect nano-size vesicles with different biochemical compositions, including liposomes synthesized with different lipid compositions, as well as EVs and lipoproteins with similar biophysical properties but dissimilar biochemical properties. Moreover, EVs derived from the same parental cells but treated with different culture conditions were characterized to investigate the correlation of impedance changes with biochemical properties and functionality in terms of pro-inflammatory responses. The system also showed the ability to discriminate between EVs derived from different cellular origins as well as among size-sorted EVs harbored from the same cellular origin. This proof-of-concept approach is the first step towards utilizing EIS as a label-free, non-invasive, and rapid sensor for detection and characterization of pathogenic EVs and other nanovesicles in the future.

A label-free and low-power microelectronic impedance spectroscopy for characterization of exosomes

“…The impedance signals were recorded at a sampling rate of 225 samples/sec and each measurement was repeated at least three times. The data analysis was obtained as previously reported by our group [26,27]. The impedance was normalized based on the 'opacity' concept to rule out the effect of the particles' number as shown in Eq (1) [28][29][30][31][32][33].…”

“…We have previously reported an insulator-based dielectrophoretic (iDEP) device to trap exosomes from biofluids by applying a low DC field across an array of micropipettes [24,25]. In addition, we have recently developed a proof-of-concept EIS system using the iDEP micropipette system with sensing electrodes manually positioned across the trapped particles by a micromanipulator [26], and a microchip comprised of microfabricated triangular posts on a substrate for entrapment of nanoscale particles using the DEP force and coplanar microelectrodes for impedance measurements of the particles [27]. The integrated microchip device could differentiate sub-micron particles including 100 nm polystyrene beads and liposomes as their impedance was measured using an AC field at 1 kHz to 10 MHz [27].…”

A Digital Microfluidic Device Integrated with Electrochemical Impedance Spectroscopy for Cell-Based Immunoassay

“…To conduct impedance measurement, the sensing electrodes were interfaced with a multiplexer (Centre Suisse d'Electronique et de Microtechnique) via a pogo pin electronic interface to connect to a multiplexer and digital impedance analyzer (HF2LI/HF2TA, Zurich Instruments, Zürich, Switzerland), which allows up to 6 analytes to be detected in parallel. An AC field with a peak amplitude of 100 mV swept from 100 Hz to 100 kHz, and frequency-based logarithmic sweep mode was used to record the amplitude and phase of the impedance signal to generate the impedance spectrum [30]. Afterward, the data was processed with a custom script written in MATLAB (MathWorks Inc., Natick, MA, USA) for statistical analysis.…”

Critical considerations in determining the surface charge of small extracellular vesicles

“…These capture molecules potentially contaminate EVs and alter their properties, thereby negatively impacting downstream analyses and their reliability for clinical applications (Gonzalez et al, 2021; Y. T. Kang et al, 2017). In contrast, focusing on the physical properties of small EVs (e.g., electrical surface charge) for the development of label-free isolation techniques supports the maintenance of native characteristics of isolated EVs along with advantages in terms of cost, time, process simplicity and reproducibility (Hassanpour Tamrin et al, 2021;Shi et al, 2019;Y. Zhang et al, 2022).…”