| 1. |
- Dev, Apurba, et al.
(author)
-
Fabrication of Periodic Nanostructure Assemblies by Interfacial Energy Driven Colloidal Lithography
- 2014
-
In: Advanced Functional Materials. - : Wiley. - 1616-301X .- 1616-3028. ; 24:29, s. 4577-4583
-
Journal article (peer-reviewed)abstract
- A novel interfacial energy driven colloidal lithography technique to fabricate periodic patterns from solution-phase is presented and the feasibility and versatility of the technique is demonstrated by fabricating periodically arranged ZnO nanowire ensembles on Si substrates. The pattern fabrication method exploits different interfaces formed by sol-gel derived ZnO seed solution on a hydrophobic Si surface covered by a monolayer of colloidal silica spheres. While the hydrophobic Si surface prevents wetting by the seed solution, the wedge shaped regions surrounding the contact point between the colloidal particles and the Si substrate trap the solution due to interfacial forces. This technique allows fabrication of uniform 2D micropatterns of ZnO seed particles on the Si substrate. A hydrothermal technique is then used to grow well-defined periodic assemblies of ZnO nanowires. Tunability is demonstrated in the dimensions of the patterns by using silica spheres with different diameters. The experimental data show that the periodic ZnO nanowire assembly suppresses the total reflectivity of bare Si by more than a factor of 2 in the wavelength range 400-1300 nm. Finite-difference time-domain simulations of the wavelength-dependent reflectivity show good qualitative agreement with the experiments. The demonstrated method is also applicable for other materials synthesized by solution chemistry.
|
|
| 2. |
- Dev Choudhury, Bikash, et al.
(author)
-
Silicon micro-structure and ZnO nanowire hierarchical assortments for light management
- 2013
-
In: Optical Materials Express. - 2159-3930 .- 2159-3930. ; 3:8, s. 1039-1048
-
Journal article (peer-reviewed)abstract
- We present fabrication and optical characterization of Si microstructure-ZnO nanowire (NWs) hierarchical structures for light management. Random and periodic hierarchical structures constituting Si micro pillar or micro pyramid arrays with overgrown ZnO NWs have been fabricated. Inexpensive colloidal lithography in combination with dry and wet chemical etching is used to fabricate Si microstructures, and ZnO NWs are grown by hydrothermal synthesis. The periodic Si micro pyramid-ZnO NWs hierarchical structure shows broadband antireflection with average reflectance as low as 2.5% in the 300-1000 nm wavelength range. A tenfold enhancement in Raman intensity is observed in this structure compared to planar Si sample. These hierarchical structures with enriched optical properties and high surface to volume ratio are promising for photovoltaic (PV) and sensor applications.
|
|
| 3. |
- Agrawal, Vipin, 1994-, et al.
(author)
-
MeMC : A package for Monte Carlo simulations of spherical shells
- 2022
-
In: Journal of Open Source Software. - : The Open Journal. - 2475-9066. ; 7:74
-
Journal article (peer-reviewed)abstract
- The MeMC is an open-source software package for Monte Carlo simulation of elastic shells. It is designed as a tool to interpret the force-distance data generated by indentation of biological nano-vesicles by atomic force microscopes. The code is written in c++ and python. The code is customizable – new modules can be added in a straightforward manner.
|
|
| 4. |
- Almeida, J. M. P., et al.
(author)
-
Femtosecond laser processing of glassy and polymeric matrices containing metals and semiconductor nanostructures
- 2013
-
In: Optical materials (Amsterdam). - : Elsevier BV. - 0925-3467 .- 1873-1252. ; 35:12, s. 2643-2648
-
Journal article (peer-reviewed)abstract
- Tailoring properties of materials by femtosecond laser processing has been proposed in the last decade as a powerful approach for technological applications, ranging from optics to biology. Although most of the research output in this field is related to femtosecond laser processing of single either organic or inorganic materials, more recently a similar approach has been proposed to develop advanced hybrid nanomaterials. Here, we report results on the use of femtosecond lasers to process hybrid nanomaterials, composed of polymeric and glassy matrices containing metal or semiconductor nanostructures. We present results on the use of femtosecond pulses to induce Cu and Ag nanoparticles in the bulk of borate and borosilicate glasses, which can be applied for a new generation of waveguides. We also report on 3D polymeric structures, fabricated by two-photon polymerization, containing Au and ZnO nanostructures, with intense two-photon fluorescent properties. The approach based on femtosecond laser processing to fabricate hybrid materials containing metal or semiconductor nanostructures is promising to be exploited for optical sensors and photonics devices.
|
|
| 5. |
- Cardoso, Marcos R., et al.
(author)
-
Highly hydrophobic hierarchical nanomicro roughness polymer surface created by stamping and laser micromachining
- 2015
-
In: Journal of Applied Polymer Science. - : Wiley. - 0021-8995 .- 1097-4628. ; 132:24
-
Journal article (peer-reviewed)abstract
- This article describes the design and fabrication of hierarchical nanomicrostructured polymer surfaces with high hydrophobicity. The nanoscale roughness is achieved by stamping a ZnO nanowire film into PDMS. Subsequently, microstructures with different periodicities are created in the stamped PDMS sample by direct laser writing using femtosecond pulses. With this approach, we were able to produce hierarchical surface morphologies, composed of nano and microscale structures that exhibit water contact angles larger than 160 degrees.
|
|
| 6. |
- Cavallaro, Sara, et al.
(author)
-
Comparison and optimization of nanoscale extracellular vesicle imaging by scanning electron microscopy for accurate size-based profiling and morphological analysis
- 2021
-
In: Nanoscale Advances. - : Royal Society of Chemistry. - 2516-0230. ; 3:11, s. 3053-3063
-
Journal article (peer-reviewed)abstract
- Nanosized extracellular vesicles (EVs) have been found to play a key role in intercellular communication, offering opportunities for both disease diagnostics and therapeutics. However, lying below the diffraction limit and also being highly heterogeneous in their size, morphology and abundance, these vesicles pose significant challenges for physical characterization. Here, we present a direct visual approach for their accurate morphological and size-based profiling by using scanning electron microscopy (SEM). To achieve that, we methodically examined various process steps and developed a protocol to improve the throughput, conformity and image quality while preserving the shape of EVs. The study was performed with small EVs (sEVs) isolated from a non-small-cell lung cancer (NSCLC) cell line as well as from human serum, and the results were compared with those obtained from nanoparticle tracking analysis (NTA). While the comparison of the sEV size distributions showed good agreement between the two methods for large sEVs (diameter > 70 nm), the microscopy based approach showed a better capacity for analyses of smaller vesicles, with higher sEV counts compared to NTA. In addition, we demonstrated the possibility of identifying non-EV particles based on size and morphological features. The study also showed process steps that can generate artifacts bearing resemblance with sEVs. The results therefore present a simple way to use a widely available microscopy tool for accurate and high throughput physical characterization of EVs.
|
|
| 7. |
- Cavallaro, Sara, 1992-
(author)
-
Development of Techniques for Characterization, Detection and Protein Profiling of Extracellular Vesicles
- 2021
-
Doctoral thesis (other academic/artistic)abstract
- Nanosized extracellular vesicles (EVs, ∼30-2000 nm) have emerged as important mediators of intercellular communication, offering opportunities for both diagnostics and therapeutics. In particular, small EVs generated from the endolysosomal pathway (∼30-150 nm), referred to as exosomes, have attracted interest as a suitable biomarker for cancer diagnostics and treatment monitoring based on minimally invasive liquid biopsies. This is because exosomes carry valuable biological information (proteins, lipids, genetic material, etc.) reflecting their cells of origin. Using EVs as biomarkers or drug delivery agents in clinical applications requires a full understanding of their cellular origin, functions, and biological relevance. However, due to their small size and very high heterogeneity in molecular and physical features, the analysis of these vesicles is challenged by the limited detection ranges and/or accuracy of the currently available techniques. To overcome some of these challenges, this thesis focuses on developing different techniques for characterization, detection and protein profiling of EVs at both bulk and single particle levels. Specifically, the three methods investigated are scanning electron microscopy, electrokinetic sensing, and combined fluorescence - atomic force microscopy. First, a protocol for scanning electron microscopy imaging of EVs was optimized to improve the throughput and image quality of the method while preserving the shape of the vesicles. Application of the developed protocol for analysis of EVs from human serum showed the possibility to use scanning electron microscopy for morphological analysis and high-resolution size-based profiling of EVs over their entire size range. Comparison with nanoparticle tracking analysis, a commonly used technique for EV size estimation, showed a superior sensitivity of scanning electron microscopy for particles smaller than 70-80 nm. Moreover, the study showed process steps that can generate artifacts resembling sEVs and ways to minimize them. Secondly, a novel label-free electrokinetic sensor based on streaming current was developed, optimized and multiplexed for EV protein analysis at a bulk level. Using multiple microcapillary sensors functionalized with antibodies, the method showed the capacity for multiplexed detection of different surface markers on small EVs from non-small-cell lung cancer cells. The device performance in the multichannel configuration remained similar to the single-channel one in terms of noise, detection sensitivity, and reproducibility. The application of the technique for analysis of EVs isolated from lung cancer patients with different genomic alterations and after different applied treatments demonstrated the prospect of using EVs from liquid biopsies as a source of biomarker for cancer monitoring. Moreover, the results held promise for the application of the developed method in clinical settings. Finally, to increase the understanding of EV subpopulations and heterogeneity, a platform combining fluorescence and atomic force microscopy was developed for multiparametric analysis of EVs at a single particle level. The use of a precise spot identification approach and an efficient vesicle capture protocol allowed to study and correlate for the first time the membrane protein composition, size and mechanical properties (Young modulus) on individual small EVs. The application of the technique to vesicles isolated from different cell lines identified both common and cell line-specific EV subpopulations bearing distinct distributions of the analyzed parameters. For example, a sEV population co-expressing all the three analyzed proteins in relatively high abundance, yet having average diameters of <100 nm and relatively low Young moduli was found in all cell lines. The obtained results highlighted the possibility of using the developed platform to help decipher unsolved questions regarding EV biology.
|
|
| 8. |
- Cavallaro, Sara, et al.
(author)
-
Label-Free Surface Protein Profiling of Extracellular Vesicles by an Electrokinetic Sensor
- 2019
-
In: ACS Sensors. - : AMER CHEMICAL SOC. - 2379-3694. ; 4:5, s. 1399-1408
-
Journal article (peer-reviewed)abstract
- Small extracellular vesicles (sEVs) generated from the endolysosomal system, often referred to as exosomes, have attracted interest as a suitable biomarker for cancer diagnostics, as they carry valuable biological information and reflect their cells of origin. Herein, we propose a simple and inexpensive electrical method for label-free detection and profiling of sEVs in the size range of exosomes. The detection method is based on the electrokinetic principle, where the change in the streaming current is monitored as the surface markers of the sEVs interact with the affinity reagents immobilized on the inner surface of a silica microcapillary. As a proof-of-concept, we detected sEVs derived from the non-small-cell lung cancer (NSCLC) cell line H1975 for a set of representative surface markers, such as epidermal growth factor receptor (EGFR), CD9, and CD63. The detection sensitivity was estimated to be similar to 175000 sEVs, which represents a sensor surface coverage of only 0.04%. We further validated the ability of the sensor to measure the expression level of a membrane protein by using sEVs displaying artificially altered expressions of EGFR and CD63, which were derived from NSCLC and human embryonic kidney (HEK) 293T cells, respectively. The analysis revealed that the changes in EGFR and CD63 expressions in sEVs can be detected with a sensitivity in the order of 10% and 3%, respectively, of their parental cell expressions. The method can be easily parallelized and combined with existing microfluidic-based EV isolation technologies, allowing for rapid detection and monitoring of sEVs for cancer diagnosis.
|
|
| 9. |
- Cavallaro, Sara, et al.
(author)
-
Multiparametric Profiling of Single Nanoscale Extracellular Vesicles by Combined Atomic Force and Fluorescence Microscopy : Correlation and Heterogeneity in Their Molecular and Biophysical Features
- 2021
-
In: Small. - : Wiley. - 1613-6810 .- 1613-6829. ; 17:14
-
Journal article (peer-reviewed)abstract
- Being a key player in intercellular communications, nanoscale extracellular vesicles (EVs) offer unique opportunities for both diagnostics and therapeutics. However, their cellular origin and functional identity remain elusive due to the high heterogeneity in their molecular and physical features. Here, for the first time, multiple EV parameters involving membrane protein composition, size and mechanical properties on single small EVs (sEVs) are simultaneously studied by combined fluorescence and atomic force microscopy. Furthermore, their correlation and heterogeneity in different cellular sources are investigated. The study, performed on sEVs derived from human embryonic kidney 293, cord blood mesenchymal stromal and human acute monocytic leukemia cell lines, identifies both common and cell line-specific sEV subpopulations bearing distinct distributions of the common tetraspanins (CD9, CD63, and CD81) and biophysical properties. Although the tetraspanin abundances of individual sEVs are independent of their sizes, the expression levels of CD9 and CD63 are strongly correlated. A sEV population co-expressing all the three tetraspanins in relatively high abundance, however, having average diameters of <100 nm and relatively low Young moduli, is also found in all cell lines. Such a multiparametric approach is expected to provide new insights regarding EV biology and functions, potentially deciphering unsolved questions in this field.
|
|
| 10. |
- Cavallaro, Sara, et al.
(author)
-
Multiplexed electrokinetic sensor for detection and therapy monitoring of extracellular vesicles from liquid biopsies of non-small-cell lung cancer patients
- 2021
-
In: Biosensors & bioelectronics. - : Elsevier. - 0956-5663 .- 1873-4235. ; 193
-
Journal article (peer-reviewed)abstract
- Liquid biopsies based on extracellular vesicles (EVs) represent a promising tool for treatment monitoring of tumors, including non-small-cell lung cancers (NSCLC). In this study, we report on a multiplexed electrokinetic sensor for surface protein profiling of EVs from clinical samples. The method detects the difference in the streaming current generated by EV binding to the surface of a functionalized microcapillary, thereby estimating the expression level of a marker. Using multiple microchannels functionalized with different antibodies in a parallel fluidic connection, we first demonstrate the capacity for simultaneous detection of multiple surface markers in small EVs (sEVs) from NSCLC cells. To investigate the prospects of liquid biopsies based on EVs, we then apply the method to profile sEVs isolated from the pleural effusion (PE) fluids of five NSCLC patients with different genomic alterations (ALK, KRAS or EGFR) and applied treatments (chemotherapy, EGFR- or ALKtyrosine kinase inhibitors). The vesicles were targeted against CD9, as well as EGFR and PD-L1, two treatment targets in NSCLC. The electrokinetic signals show detection of these markers on sEVs, highlighting distinct interpatient differences, e.g., increased EGFR levels in sEVs from a patient with EGFR mutation as compared to an ALK-fusion one. The sensors also detect differences in PD-L1 expressions. The analysis of sEVs from a patient prior and post ALK-TKI crizotinib treatment reveals significant increases in the expressions of some markers (EGFR and PD-L1). These results hold promise for the application of the method for tumor treatment monitoring based on sEVs from patient liquid biopsies.
|
|
