Reliable Detection of Extracellular PD-L1 by DNA Computation-Mediated Microfluidics

Abstract

Extracellular vesicle PD-L1 (programmed death-1 ligand 1) is of greater value in tumor diagnosis, prognosis, and efficacy monitoring of anti-PD-1/PD-L1 immunotherapy. However, soluble PD-L1 interferes with the accurate detection of extracellular vesicle (EV) PD-L1. Here, we developed a microfluidic differentiation method for the detection of extracellular PD-L1, without the interference of soluble, by DNA computation with lipid probes and PD-L1 aptamer as inputs (DECLA). For the developed DECLA method, a cholesterol-DNA probe was designed that efficiently embeds into the EV membrane, and an aptamer-based PD-L1 probe was used for PD-L1 recognition. Due to the stable secondary structure of the designed connector, only cobinding of cholesterol-DNA and PD-L1 affinity probe induced biotin-labeled connector activation, while soluble PD-L1 cannot hybridize. As a result, PD-L1 EVs can be efficiently captured by streptavidin-functioned herringbone chip and quantified by anti-CD63-induced fluorescence signal. The high specificity of dual-input DNA computation allied to the high sensitivity of microfluidic-based detection was suitable for distinguishing lung cancer patients from healthy donors, highlighting its potential translation to clinical diagnosis and therapy monitoring.

Electroosmotic Micromixing in Physicochemically Patterned Microchannels

“…It is now well-known that there are multifold advantages of using microfluidic devices over macroscopic ones owing to their portability, ease of use, availability of a higher surface-to-volume ratio for the process intensified engineering processes, control over the reagent parameters owing to their usage of smaller volumes, and capacity to bring in the aspects of very-large-scale integration (VLSI) for a larger throughput and multitasking, among others. Thus, such microfluidic platforms are found to appear in diverse modern-day functionalities that include drug delivery, − point-of-care diagnostics, − tissue engineering, − high-throughput screening, − protein crystallization, − and deoxyribonucleic acid (DNA) analysis. , In particular, the success in the integration of multiplexing of microfluidic devices on the lab-on-a-chip , platforms has led to the development of portable laboratory prototypes in the diverse areas of biology, − chemistry, − medicine, , and engineering. , However, several limitations related to microfluidic platforms have emerged over the years, including the diffusion-limited mixing capacity, a relatively lower throughput, and crowding–clogging of transport materials, among others. Of late, such areas have become intense scientific and engineering research.…”

Engineering Cell Membranes: From Extraction Strategies to Emerging Biosensing Applications

“…22 molecules for cell−cell communication. 23,24 The method employed for exosomes extraction is somewhat similar to that used to extract living cell membranes. 25 In terms of applications, both can be used in biomolecular detection and drug delivery.…”

“…Generally, except for plasma membranes in the typical sense, extracted cell membranes also cover organelle membranes and exosomes . Exosomes are nanosized extracellular vesicles secreted by cells, delivering important biologically active molecules for cell–cell communication. , The method employed for exosomes extraction is somewhat similar to that used to extract living cell membranes . In terms of applications, both can be used in biomolecular detection and drug delivery.…”

Accurate Cancer Screening and Prediction of PD-L1-Guided Immunotherapy Efficacy Using Quantum Dot Nanosphere Self-Assembly and Machine Learning

“…Gene sequencing techniques, such as real-time quantitative fluorescence polymerase chain reaction (qPCR) 14 and isothermal amplification, 15 are used to detect PD-L1 gene expression levels to indirectly reflect the expression of the PD-L1 protein. Despite the great success of the above studies in quantitative PD-L1 detection, 16,17 these methods often require complex experimental steps and expensive equipment, and reliable methods for cancer screening and quantitative analysis of PD-L1 expression levels by circulating exosomes are still less reported. Nanomaterial-assisted signal amplification is a method of using the properties of nanomaterials to enhance the signal through the multieffective and precise assembly of nanomaterials to achieve cyclic amplification of the signal, thus improving the detection sensitivity.…”

“…Gene sequencing techniques, such as real-time quantitative fluorescence polymerase chain reaction (qPCR) and isothermal amplification, are used to detect PD-L1 gene expression levels to indirectly reflect the expression of the PD-L1 protein. Despite the great success of the above studies in quantitative PD-L1 detection, , these methods often require complex experimental steps and expensive equipment, and reliable methods for cancer screening and quantitative analysis of PD-L1 expression levels by circulating exosomes are still less reported.…”