Owing to their excellent multiplexing ability, high sensitivity, and large dynamic range, immunoassays using surface-enhanced Raman scattering (SERS) as the readout signal have found prosperous applications in fields such as disease diagnosis, environmental surveillance, and food safety supervision. Various ever-increasing demands have promoted SERS-based immunoassays from the classical sandwich-type ones to those integrated with fascinating automatic platforms (e.g., test strips and microfluidic chips). As recent years have witnessed impressive progress in SERS immunoassays, we try to comprehensively cover SERS-based immunoassays from their basic working principles to specific applications. Focusing on several basic elements in SERS immunoassays, typical structures of SERS nanoprobes, productive optical spectral encoding strategies, and popular immunoassay platforms are highlighted, followed by their representative biological applications in the last 5 years. Moreover, despite the vast advances achieved ...

SERS-Activated Platforms for Immunoassay: Probes, Encoding Methods, and Applications

Extracellular vesicles (EVs) are a family of small membrane vesicles that carry information about cells by which they are secreted. Growing interest in the role of EVs in intercellular communication, but also in using their diagnostic, prognostic and therapeutic potential in (bio) medical applications, demands for accurate assessment of their biochemical and physical properties. In this review, we provide an overview of available technologies for EV analysis by describing their working principles, assessing their utility in EV research and summarising their potential and limitations. To emphasise the innovations in EV analysis, we also highlight the unique possibilities of emerging technologies with high potential for further development.

Extracellular Vesicle Quantification and Characterization: Common Methods and Emerging Approaches

A visible and colorimetric aptasensor based on DNA-capped single-walled carbon nanotubes for detection of exosomes.

As a kind of most important cancer biomarker, exosomes are getting more frequently investigated in cancer diagnosis. In this study, we proposed an SERS-based method for the screening and simultaneous multiple detection of exosomes using magnetic substrates and SERS probes. Specifically, the capturing substrates are achieved using gold shell magnetic nanobeads modified by aptamers, which can capture most kinds of exosomes by recognizing the generic surface protein CD63. Moreover, the SERS probes are made of gold nanoparticles decorated with a Raman reporter and a specific aptamer for targeting exosomes. Further, for the simultaneous detection of multiple kinds of exosomes, three kinds of SERS probes were designed using different SERS reporters. While detecting specific kinds of exosomes, the capturing substrates were mixed with these three kinds of SERS probes. When the target exosome is present, an apta-immunocomplex can be formed among the target exosomes, the substrate, and the corresponding kind of SERS probes, and the other non-specific SERS probes remain in the suspension. Hence, an SERS signal with a decreased intensity will be detected in the supernatant, indicating the presence of the target exosomes. Finally, this detection method has also been successfully employed for the detection of exosomes in real blood samples; this proves that the proposed SERS-based method is a promising tool for clinical cancer screening based on exosomes.

Screening and multiple detection of cancer exosomes using an SERS-based method.

The detection of exosomes is promising for the early diagnosis of cancer. However, the development of suitable optical sensors remains challenging. We have developed the first luminescent nanosensor for the multiplex differentiation of cancer exosomes that bypasses real-time light excitation. The sensor is composed of a near-infrared semiconducting polyelectrolyte (ASPN) that forms a complex with a quencher-tagged aptamer. The afterglow signal of the nanocomplex (ASPNC), being initially quenched, is turned on in the presence of aptamer-targeted exosomes. Because detection of the afterglow takes place after the excitation, background signals are minimized, leading to an improved limit of detection that is nearly two orders of magnitude lower than that of fluorescence detection in cell culture media. Also, ASPNC can be easily tailored to detect different exosomal proteins by changing the aptamer sequence. This enables an orthogonal analysis of multiple exosome samples, potentially permitting an accurate identification of the cellular origin of exosomes for cancer diagnosis.

/pdf/near-infrared-afterglow-semiconducting-nano-polycomplexes-mg8jxmbvrc.pdf

Near-Infrared Afterglow Semiconducting Nano-Polycomplexes for the Multiplex Differentiation of Cancer Exosomes.

Exosomes play an important role in intercellular communications. Here, we present a surface-enhanced Raman scattering (SERS) based detection method for tumor-derived exosomes using SERS nanoprobes and magnetic nanobeads. The SERS nanoprobe has a core–shell structure, with gold core–silver shell nanorods (Au@Ag NRs) as the SERS active core, Raman molecules as the SERS reporter and a silica layer as the protecting shell. The outmost surface of the SERS nanoprobe is decorated with exosome-specific antibodies. Each magnetic nanobead is fabricated by coating Fe3O4 nanoparticles (NPs) with a silica shell and attaching specific antibodies to the silica shell. With target exosomes present, the magnetic nanobeads and SERS nanoprobes can capture the exosomes by forming a sandwich-type immunocomplex. The immunocomplex (as well as the SERS nanoprobes) can be precipitated with a magnet, and thus SERS signals can be detected in the precipitates. With no target exosomes present, no immunocomplex can be formed, and thus quite weak SERS signals will be detected in the precipitates. Hence, the SERS signal of the final magnetic separation product can be used to detect exosomes. In the experiment, using one kind of tumor cells and one kind of normal cells, we proved that the presented method can be used for both qualitative and quantitative detection of tumor-derived exosomes.

Facile detection of tumor-derived exosomes using magnetic nanobeads and SERS nanoprobes

Assessing telomere length is of vital importance since telomere length is closely related with several fatal diseases such as atherosclerosis and cancer. Here, we present a strategy to assess/measure telomere length, that is, surface enhanced Raman scattering (SERS) based in situ hybridization (SISH). The SISH method uses two kinds of SERS nanoprobes to hybridize in situ with telomeres and centromeres, respectively. The telomere specific SERS nanoprobe is called the Telo-probe, while the centromere specific SERS nanoprobe is called the Centro-probe. They are composed of metal nanoparticles (NPs), Raman reporter molecules and specially designed DNA strands. With longer telomeres, more Telo-probes will hybridize with them, resulting in a stronger SERS signal. To exclude possible influence of the SERS intensity by external factors (such as the nanoprobe concentration, the cell number or different batches of nanoprobes), centromeres are used as the inner control, which can be recognized by Centro-probes. Telo...

Surface Enhanced Raman Scattering Based in Situ Hybridization Strategy for Telomere Length Assessment

The recent proposition to combine liposomes with nanoparticles presents great opportunities to develop multifunctional drug delivery platforms. Although impressive progress has been made, attempts to elucidate the role nanoparticles play in the integral nanohybrids are still rather limited. Here, using surface-enhanced Raman scattering (SERS) technique, we investigate the influence of metal nanoparticles on the liposomal properties, ranging from drug release to intracellular movement. Specifically, we prepared SERS-active nanohybrids by attaching metal nanoparticles to liposomes and employed SERS signals to explore the intracellular behavior of the nanohybrids. Once deposited on the cell membrane, the nanohybrids entered tumor cells via clathrin-mediated endocytosis and then moved to lysosomes. In comparison with pure liposomes, metal nanoparticles in the nanohybrids had little influence on the properties of liposomes. This study fills the gap of the function of nanoparticles in the overall nanohybrids, which provides a significant prerequisite for efficient drug delivery in therapeutic applications.

Investigating the Intracellular Behaviors of Liposomal Nanohybrids via SERS: Insights into the Influence of Metal Nanoparticles.

Single molecule localization based super resolution imaging techniques (e.g. PALM/STORM) require that the employed optical nanoprobes possess signal fluctuations under certain excitations. Here, we present a Forster resonance energy transfer (FRET) based dual emission nanoprobe (denoted as FREDEN), which is suitable for PALM/STORM imaging. Basically, FREDEN is constructed by attaching Alexa Fluor 647 (A647) molecules onto semiconductor quantum dots (QDs). Dual emission is realized via FRET from QDs to A647. Experimental results confirmed that under 405 nm excitation, FREDEN exhibits two distinctive fluorescence emission bands centered at around 610 nm and 670 nm. Besides, FREDEN shows improved fluorescence fluctuation (“blinking”) behavior at the two emission bands as compared with solitary QDs or A647. Moreover, fluorescence blinking of FREDEN requires no special buffer solution, which makes the presented nanoprobe an excellent candidate for live cell PALM/STORM imaging. Using FREDEN, dual color PALM/STORM imaging of live cells is realized under a single excitation wavelength (i.e. 405 nm). With the improved fluorescence blinking behavior, the presented FREDEN shows great potential in simplifying and promoting single molecule localization based super resolution imaging.

A FRET based dual emission nanoprobe (FREDEN) with improved blinking behavior for single molecule localization imaging

The invention discloses a method for quickly detecting exosomes through an SERS (Surface Enhanced Raman Scattering) signal. The method comprises the following steps: firstly, identifying the exosomes in a sample through an immune response by utilizing a magnetic capture substrate and a Raman microprobe to form a sandwich structure in the form of magnetic capture substrate-exosome-Raman microprobe; then, separating a collected product by utilizing an externally applied magnetic field; finally, analyzing the SERS signal of the product by utilizing a confocal Raman microscopy system, wherein the SERS signal is stronger, and the concentration of the exosomes is higher. According to the method, the SERS technology is applied to detecting the exosomes; precise qualitative and quantitative analysis is performed on the sample by adopting the SERS technology, and quick and high-sensitivity detection of the exosomes can be realized.

Method for quickly detecting exosomes through SERS signal

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