Ultrasensitive detection of circulating exosomes with a 3D-nanopatterned microfluidic chip

Abstract

The performance of current microfluidic methods for exosome detection is constrained by boundary conditions and by fundamental limits to microscale mass transfer and to interfacial exosome binding. Here, we show that a microfluidic chip designed with self-assembled 3D herringbone nanopatterns can detect low levels of tumour-associated exosomes in plasma (10 exosomes μL −1 , or approximately 200 vesicles per 20-μL spiked sample) that would otherwise be undetectable by standard microfluidic systems for biosensing. The nanopatterns promote microscale mass transfer, increase surface area and probe density to enhance the efficiency and speed of exosome binding, and permit drainage of the boundary fluid to reduce near-surface hydrodynamic resistance, thus promoting particle–surface interactions for exosome binding. We used the device for the detection, in 2-μL plasma samples from 20 ovarian cancer patients and from 10 age-matched controls, of exosome subpopulations expressing CD24, EpCAM, and FRalpha proteins, and suggest exosomal FRalpha as a potential biomarker for the early detection and progression monitoring of ovarian cancer. The nanolithography-free nanopatterned device should facilitate the use of liquid biopsies for cancer diagnosis.

Homogeneous, Low‐volume, Efficient, and Sensitive Quantitation of Circulating Exosomal PD‐L1 for Cancer Diagnosis and Immunotherapy Response Prediction

“…[6] Thus,exosomal PD-L1 can be applied as an ideal complement indicator for the existing immune checkpoint diagnosis,w ith the advantages of accuracy,n on-invasiveness,a nd real-time monitoring. [7] Although some methods of exosome analysis based on biosensing [8] and microfluidic technology [9] have been reported, these methods have not been applied to the quantification of exosomal PD-L1 in real clinical samples. Theexosomal PD-L1 quantitation reported so far is based on the well-established ELISA.…”

Isolation and Profiling of Circulating Tumor‐Associated Exosomes Using Extracellular Vesicular Lipid–Protein Binding Affinity Based Microfluidic Device

“…DiO Staining of the Extracellular Vesicles : Staining of extracellular vesicles using lipophilic dyes such as DiO and PKH has been used in various studies . DiO staining was conducted in two experiments.…”

“…Microfluidics technologies offer many advantages and may become the optimal method for exosome isolation in the future. Owing to recent advances in microfluidic technologies, several microfluidic devices for exosome isolation have been developed with better recovery and shorter processing times compared to UC . Among them, immunoaffinity‐based microfluidic isolation using antibodies against exosomal surface proteins is advantageous as it allows for high specificity exosome isolation from heterogeneous samples, such as plasma, serum, or urine.…”

“…Among them, immunoaffinity‐based microfluidic isolation using antibodies against exosomal surface proteins is advantageous as it allows for high specificity exosome isolation from heterogeneous samples, such as plasma, serum, or urine. This method also has been incorporated with an engineered surface or characteristic patterns in order to enhance the efficiency of exosome isolation . As an example, antibodies against the tetraspanin CD63 have been widely applied to exosome isolation from the plasma of patients with ovarian cancer, breast cancer, and glioblastoma .…”

“…This method also has been incorporated with an engineered surface or characteristic patterns in order to enhance the efficiency of exosome isolation . As an example, antibodies against the tetraspanin CD63 have been widely applied to exosome isolation from the plasma of patients with ovarian cancer, breast cancer, and glioblastoma . However, anti‐CD63 is not a specific biomarker for any one cancer, and its expression is known to vary depending on the type of cancer .…”