Quantitative interferometric reflectance imaging for the detection and measurement of biological nanoparticles

Sevenler, Derin; Avcı, Oğuzhan; Ünlü, M. Selim

doi:10.1364/boe.8.002976

Cited by 45 publications

(58 citation statements)

References 37 publications

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“…If this interpretation is correct, then WT CD63containing exosomes should be the same size as CD63/Y235A-containing exosomes. To test this prediction, we used single-particle interferometry reflectance (SPIR) imaging (Avci et al, 2015;Daaboul et al, 2017;Daaboul et al, 2016;Sevenler et al, 2017;Sevenler et al, 2018) and immunofluorescence microscopy (IFM) to measure the sizes of thousands of WT CD63 exosomes and CD63/Y235A exosomes. SPIRI-IFM is a novel analytical approach that combines traditional IFM of immunolabeled exosomes with label-free visualization of exosomes using a single-particle interferometric reflectance imaging sensor, in which the interference of light reflected from the sensor surface is (i) modified by the presence of a chip-bound exosome, (ii) varies in relation to the diameter of the chip-bound exosome, and (iii) allows measurement of exosome diameter to a resolution of 0.5 nm.…”

Section: Resultsmentioning

confidence: 99%

A shared pathway of exosome biogenesis operates at plasma and endosome membranes

Fordjour

Daaboul

Gould

2019

Preprint

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This study of exosome cargo protein budding reveals that cells use a common pathway for budding exosomes from plasma and endosome membranes, providing a new mechanistic explanation for exosome heterogeneity and a rational roadmap for exosome engineering. AbstractEukaryotic cells secrete exosomes, which are small (~30-200 nm dia.), single membranebound organelles that transmit signals and molecules to other cells. Exosome-mediated signaling contributes to diverse physiological and disease processes, rendering their biogenesis of high biomedical importance. The prevailing hypothesis is that exosomes bud exclusively at endosome membranes and are released only upon endosome fusion with the plasma membrane. Here we tested this hypothesis by examining the intracellular sorting and exosomal secretion of the exosome cargo proteins CD63, CD9, and CD81. We report here that CD9 and CD81 are both localized to the plasma membrane and bud >5-fold more efficiently than endosome-localized CD63. Furthermore, we show that redirecting CD63 from endosomes to the plasma membrane by mutating its endocytosis signal (CD63/Y235A) increased its exosomal secretion ~6-fold, whereas redirecting CD9 to endosomes by adding an endosome targeting signal (CD9/YEVM) reduced its exosomal secretion ~5-fold. These data demonstrate that the plasma membrane is a major site of exosome biogenesis, and more importantly, that cells possess a common pathway for exosome protein budding that operates at both plasma and endosome membranes. Using a combination of single-particle interferometry reflectance (SPIR) imaging and immunofluorescence (IF) microscopy, we also show that variations in exosome composition are controlled by differential intracellular protein trafficking rather than by separate mechanisms of exosome biogenesis. This new view of exosome biogenesis offers a simple explanation for the pronounced compositional heterogeneity of exosomes and a validated roadmap for exosome engineering.

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Section: Resultsmentioning

confidence: 99%

A shared pathway of exosome biogenesis operates at plasma and endosome membranes

Fordjour

Daaboul

Gould

2019

Preprint

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“…Extension of the scheme to multiport interferometry [31] and the associated phase super-resolution may make possible imaging of previously inaccessible smaller bodies such as exoplanets, moons and asteroids. In biology and medical imaging-through incorporation into interferometric reflectance schemes that detect reflected thermal light shone onto the source [32]the Count and Click schemes can provide optimal imaging of small biological entities. B.…”

Section: Arxiv:181102192v3 [Quant-ph] 17 Jul 2019mentioning

confidence: 99%

Optimal Imaging of Remote Bodies Using Quantum Detectors

et al. 2019

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We implement a general imaging method by measuring the complex degree of coherence using linear optics and photon number resolving detectors. In the absence of collective or entanglementassisted measurements, our method is optimal over a large range of practically relevant values of the complex degree of coherence. We measure the size and position of a small distant source of pseudo-thermal light, and show that our method outperforms the traditional imaging method by an order of magnitude in precision. Finally, we show that a lack of photon number resolution in the detectors has only a modest detrimental effect on measurement precision and simulate imaging using the new and traditional methods with an array of detectors; showing that the new method improves both image clarity and contrast.

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“…The oxide film thickness of the substrate, illumination wavelength and nanorod geometry all effect the amplitude of the scattered light collected by the objective. We used a quantitative model of interferometric reflectance imaging described previously to co-optimize these various parameters and guide the selection of 25 nm diameter GNRs with a longitudinal surface plasmon resonance wavelength of 650 nm, a substrate oxide thickness of 110 nm, and 650 nm LED illumination source with an 10 nm FWHM bandpass filter ( Figure S2) 27 . Early on in this study, we also compared the circular polarization scheme described here with a simpler cross polarization scheme, in which the illumination is linearly polarized rather than circularly polarized, and mostly blocked by a crossed polarizer in the collection path ( Figure S3).…”

Section: Resultsmentioning

confidence: 99%

Digital Microarrays: Single-Molecule Readout with Interferometric Detection of Plasmonic Nanorod Labels

et al. 2018

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DNA and protein microarrays are a high-throughput technology that allow the simultaneous quantification of tens of thousands of different biomolecular species. The mediocre sensitivity and limited dynamic range of traditional fluorescence microarrays compared to other detection techniques have been the technology's Achilles' heel and prevented their adoption for many biomedical and clinical diagnostic applications. Previous work to enhance the sensitivity of microarray readout to the single-molecule ("digital") regime have either required signal amplifying chemistry or sacrificed throughput, nixing the platform's primary advantages. Here, we report the development of a digital microarray which extends both the sensitivity and dynamic range of microarrays by about 3 orders of magnitude. This technique uses functionalized gold nanorods as single-molecule labels and an interferometric scanner which can rapidly enumerate individual nanorods by imaging them with a 10× objective lens. This approach does not require any chemical signal enhancement such as silver deposition and scans arrays with a throughput similar to commercial fluorescence scanners. By combining single-nanoparticle enumeration and ensemble measurements of spots when the particles are very dense, this system achieves a dynamic range of about 6 orders of magnitude directly from a single scan. As a proof-of-concept digital protein microarray assay, we demonstrated detection of hepatitis B virus surface antigen in buffer with a limit of detection of 3.2 pg/mL. More broadly, the technique's simplicity and high-throughput nature make digital microarrays a flexible platform technology with a wide range of potential applications in biomedical research and clinical diagnostics.

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Quantitative interferometric reflectance imaging for the detection and measurement of biological nanoparticles

Cited by 45 publications

References 37 publications

A shared pathway of exosome biogenesis operates at plasma and endosome membranes

A shared pathway of exosome biogenesis operates at plasma and endosome membranes

Optimal Imaging of Remote Bodies Using Quantum Detectors

Digital Microarrays: Single-Molecule Readout with Interferometric Detection of Plasmonic Nanorod Labels

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