Focusing light deep inside living tissue has not been achieved despite its promise to play a central role in biomedical imaging, optical manipulation and therapy. To address this challenge, ...internal-guide-star-based wavefront engineering techniques--for example, time-reversed ultrasonically encoded (TRUE) optical focusing--were developed. The speeds of these techniques, however, were limited to no greater than 1 Hz, preventing them from in vivo applications. Here we improve the speed of optical focusing deep inside scattering media by two orders of magnitude, and focus diffuse light inside a dynamic scattering medium having a speckle correlation time as short as 5.6 ms, typical of living tissue. By imaging a target, we demonstrate the first focusing of diffuse light inside a dynamic scattering medium containing living tissue. Since the achieved focusing speed approaches the tissue decorrelation rate, this work is an important step towards in vivo deep tissue noninvasive optical imaging, optogenetics and photodynamic therapy.
Gold nanoflowers (GNFs) exhibit stronger light scattering ability than gold nanospheres (GNSs) with the same diameter, thereby contributing to enhancing the sensitivity of the scattering-based ...sensing method. However, the application of GNFs in biosensors based on dynamic light scattering (DLS) has not been yet reported. Herein, we describe for the first time an improved no-wash immunosensor based on dynamic light scattering for the detection of
O157:H7 (
O157:H7) in milk using GNFs for sensitive signal transduction. To achieve this goal, a thiolated amphiphilic carboxyl ligand was introduced to modify the GNF surface and improve solution stability and antibody functionalization. Several key factors that affect the detection sensitivity of our developed GNF_DLS immunosensor were systematically investigated. Under the optimal conditions, our proposed GNF_DLS immunosensor provided an excellent linear detection for
O157:H7 within the range from 6 × 10
to 6 × 10
colony-forming units (CFU)/mL, with a limit of detection of 2.7 CFU/mL. Combined with our previously reported two-step large-volume immunomagnetic separation (IMS) method, the designed GNF_DLS immunosensor can sensitively, selectively, and accurately detect the presence of
O157:H7 in pasteurized milk. The potential of our GNF_DLS method for monitoring the presence of a single bacterial cell in 1 mL of sample solution was also demonstrated. Overall, the developed GNF_DLS immunosensor can be used for the rapid and high-sensitivity determination of pathogenic bacteria and can be extended for the ultrasensitive no-wash detection of other trace analytes.
•AFM, SEM; TEM and DLS all compared.•Silica, gold, and polymer nanoparticles characterised.•AFM and TEM shown to be most appropriate for small particles.•SEM just as accurate as AFM and TEM for ...larger particles.•DLS shows dynamic behaviour but cannot characterise mixtures.
Nanoparticles have properties that depend critically on their dimensions. There are a large number of methods that are commonly used to characterize these dimensions, but there is no clear consensus on which method is most appropriate for different types of nanoparticles.
In this work four different characterization methods that are commonly applied to characterize the dimensions of nanoparticles either in solution or dried from solution are critically compared. Namely, transmission electron microscopy (TEM), scanning electron microscopy (SEM), atomic force microscopy (AFM), and dynamic light scattering (DLS) are compared with one another. The accuracy and precision of the four methods applied nanoparticles of different sizes composed of three different core materials, namely gold, silica, and polystyrene are determined. The suitability of the techniques to discriminate different populations of these nanoparticles in mixtures are also studied.
The results indicate that in general, scanning electron microscopy is suitable for large nanoparticles (above 50 nm in diameter), while AFM and TEM can also give accurate results with smaller nanoparticles. DLS reveals details about the particles’ solution dynamics, but is inappropriate for polydisperse samples, or mixtures of differently sized samples. SEM was also found to be more suitable to metallic particles, compared to oxide-based and polymeric nanoparticles. The conclusions drawn from the data in this paper can help nanoparticle researchers choose the most appropriate technique to characterize the dimensions of nanoparticle samples.
Particle agglomeration is relevant to numerous industrial applications and consumer products. The present work explores interactions between and agglomeration of gamma (γ)-alumina nanoparticles in ...pure water and dilute aqueous salt solutions. To characterize surface- and salt-specific effects, potential of mean force (PMF) profiles between γ-alumina surfaces (110 and 100 facets) are extracted using classical molecular dynamics (MD) simulations. Supporting experiments are conducted using dynamic light scattering (DLS) to investigate agglomeration at the macroscale. The ion pairs considered are sodium chloride, ammonium acetate, barium nitrate, and barium acetate; sampling a broad range of the Hofmeister series. As particle surfaces approach contact, free-energy fluctuations of the PMF profiles reflect structural adjustments of the intervening aqueous phase. We extract values for the cohesive energy from the MD results, and parse the resultant effective pair interactions into van der Waals and electrostatic contributions. Molecular scale findings from simulations correlate with hydrodynamic radii of γ-alumina nanoparticles, obtained from DLS experiments. The results highlight the applicability of molecular simulations to identify the origins of macroscale observables.
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Exosomes are cell-secreted nanovesicles present in biological fluids in normal and diseased conditions. Owing to their seminal role in cell-cell communication, emerging evidences suggest that ...exosomes are fundamental regulators of various diseases. Due to their potential usefulness in disease diagnosis, robust isolation and characterization of exosomes is critical in developing exosome-based assays. In the last few years, different exosome characterization methods, both biophysical and molecular, have been developed to characterize these tiny vesicles. Here, in this review we summarize: first, biophysical techniques based on spectroscopy (e.g., Raman spectroscopy, dynamic light scattering) and other principles, for example, scanning electron microscopy, atomic force microscopy; second, antibody-based molecular techniques including flow cytometry, transmission electron microscopy and third, nanotechnology-dependent exosome characterization methodologies.
Viruses are increasingly used as vectors for delivery of genetic material for gene therapy and vaccine applications. Recombinant adeno-associated viruses (rAAVs) are a class of viral vector that is ...being investigated intensively in the development of gene therapies. To develop efficient rAAV therapies produced through controlled and economical manufacturing processes, multiple challenges need to be addressed starting from viral capsid design through identification of optimal process and formulation conditions to comprehensive quality control. Addressing these challenges requires fit-for-purpose analytics for extensive characterization of rAAV samples including measurements of capsid or particle titer, percentage of full rAAV particles, particle size, aggregate formation, thermal stability, genome release, and capsid charge, all of which may impact critical quality attributes of the final product. Importantly, there is a need for rapid analytical solutions not relying on the use of dedicated reagents and costly reference standards. In this study, we evaluate the capabilities of dynamic light scattering, multiangle dynamic light scattering, and SEC-MALS for analyses of rAAV5 samples in a broad range of viral concentrations (titers) at different levels of genome loading, sample heterogeneity, and sample conditions. The study shows that DLS and MADLS
can be used to determine the size of full and empty rAAV5 (27 ± 0.3 and 33 ± 0.4 nm, respectively). A linear range for rAAV5 size and titer determination with MADLS was established to be 4.4 × 10
-8.7 × 10
cp/mL for the nominally full rAAV5 samples and 3.4 × 10
-7 × 10
cp/mL for the nominally empty rAAV5 samples with 3-8% and 10-37% CV for the full and empty rAAV5 samples, respectively. The structural stability and viral load release were also inferred from a combination of DLS, SEC-MALS, and DSC. The structural characteristics of the rAAV5 start to change from 40 °C onward, with increasing aggregation observed. With this study, we explored and demonstrated the applicability and value of orthogonal and complementary label-free technologies for enhanced serotype-independent characterization of key properties and stability profiles of rAAV5 samples.
Hydrodynamic and light scattering methods are urgently required for accurate characterization of nanoparticles (NPs) in the field of nanomedicine to unveil their sizes and distributions. A ...fundamental characterization approach in the field of nanomedicines is, next to standard batch dynamic light scattering (DLS) and increasingly more applied (asymmetrical flow) field-flow fractionation (FFF) coupled to multi-angle laser light scattering (MALLS), the utilization of an analytical ultracentrifuge (AUC). Here, we demonstrate the power of an AUC in comparison to batch DLS and FFF-MALLS to decipher, in detail, the size and dispersity of pharma-relevant, commercial and in-house prepared soft matter NPs, suitable for life science applications. In this study, size and dispersity of poly(lactic-co-glycolic acid) (PLGA) NPs and in-house prepared NPs, consisting of the commercially available pharmapolymer Eudragit® E or of a polymer of similar composition synthesized via reversible addition fragmentation chain transfer (RAFT) polymerization, were investigated. Simultaneously, an insight on the presence of the utilized surfactant on the NP formulations, which is usually limited with other techniques, could be achieved by multi-speed experiments with the AUC in one experimental setting. While the repeatability and ruggedness of observations with modern AUC instruments of the newest generation is demonstrated, the results are further underpinned by the classical relations of hydrodynamics. Investigations aiming at hydrodynamic diameters (from DLS) and radii of gyration (from FFF-MALLS) are critically discussed and compared to the repeatable and rugged investigations by an AUC. The latter is proven to provide a self-sufficient experimental approach for NP characterization in the field of nanomedicine based on absolute principles, compares well to FFF-MALLS, and can unravel issues in NP sizing that arise when more common techniques, such as DLS, are used.
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•Medical applications of nanoparticles (NPs) are dependent on their sizes and charge.•Determination of accurate NP size and dispersity are urgently required.•Comparison of high-end hydrodynamic, separation, and light scattering techniques.•Ruggedness and repeatability of analytical ultracentrifuge experiments.•Limits of standard and advanced techniques are discussed on par to each other.
Abstract After briefly introducing the theoretical equations for DLS based particle size analysis, the need for angular dependent DLS investigations is emphasized to obtain correct particle sizes. ...Practical examples are given that demonstrate the possible magnitudes of errors in particle size if DLS is measured at one large scattering angle, only, as done by essentially all, most frequently utilized commercial “single angle” particle sizers. The second part is focused on a novel DLS application to sensitively trace (nano)particle interactions with concentrated blood serum or plasma that leads to the formation of large aggregates in a size regime of ≫100 nm. Most likely, such aggregates originate from protein induced bridging of nanoparticles, since it is well known that serum proteins adsorb onto the surface of essentially all nanoparticles utilized in medical applications. Thus, the protein corona around nanoparticles does not only change their biological identity but to a large extend also their size, thus possibly affecting biodistribution and in vivo circulation time.
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