142 research outputs found
Need for Speed: Imaging Biological Ultrastructure with the 64-beams FAST-EM
No full textGreen Open Access added to TU Delft Institutional Repository 'You share, we take care!' - Taverne project https://www.openaccess.nl/en/you-share-we-take-care Otherwise as indicated in the copyright section: the publisher is the copyright holder of this work and the author uses the Dutch legislation to make this work public.ImPhys/Hoogenboom grou
Electron Beam-Induced Fluorescence Localization : Implementation and Feasibility in Integrated Light-Electron Microscopy
No full textCorrelative light-electron microscopy (CLEM) combines the molecular specificity of fluorescence microscopy (FM) with the ultrastructural resolution of electron microscopy (EM) to provide functional information in the context of structural detail. However, the correlation between the two modalities is hindered by a 100-fold resolution gap. Superresolution fluorescence microscopy (SR-FM) has enabled more accurate correlation in CLEM, aided by advancements in sample preparation, bimodal registration, and optimized workflows. As the resolution of FM approaches that of EM, SR-FM becomes increasingly challenging due to the need for accurate registration and preservation of fluorophore properties during EM sample preparation. Integrated microscopes can remove the need for external alignment markers and achieve high registration accuracies, eliminating sample deformations and facilitating SR-CLEM. Yet, this necessitates samples that are simultaneously amenable to both FM and EM. While advancements are being made to engineer fixation-resistant fluorescent proteins and develop preparation protocols for preserving in-resin fluorescence, it is worth exploring the possibilities of the unified platform that integrated light-electron microscopy offers, especially for SR-FM. In caseswhere traditional SR-FM cannot be used due to vacuum or other limitations, the combination of both light and electron microscopy can provide valuable multi-modal information. It also enables central control of the experimental system, thereby offering new ways to manipulate, process, and interpret fluorescence data. This thesis aims to investigate and utilize luorescent response to electron irradiation using integrated light-electronmicroscopy....ImPhys/Hoogenboom grou
Cathodoluminescence Microscopy of Nanostructures on Transparent Substrates
No full textCathodoluminescence (CL), the excitation of light by an electron beam, has gained attention as an analysis tool for investigating the optical response of a structure, at a resolution that approaches that in electron microscopy, in the nanometer range. However, the application possibilities are limited because the use of transparent substrates, one of the most common sample substrates for optical characterizations in multiple research fields, is normally avoided in CL microscopy, since these materials generate a strong signal that contributes as a background to the measurement. The main goal in this thesis is to achieve cathodoluminescence detection of nanostructures on glass-based substrates. For that purpose, a CL system with enhanced collection efficiency and confocal detection of the signal was developed, built and tested. The design is based on an integrated Scanning Electron and Optical Microscope, a setup that offers simultaneous correlated acquisition of the electron and light signals. Besides cathodoluminescence, other interesting applications derive from the combination of these techniques, but they are out of the scope of this thesis. Chapter 1 intends to give general introduction to cathodoluminescence as a microscopy analysis tool. First we discuss its generation principle: considering the excitation and emission mechanisms, electron-hole recombination, transition radiation and surface plasmon polariton radiative outcoupling are identified as the main CL sources in the structures investigated in this thesis. In bulk samples, the emission is not restricted to the nanometer size spot where the incoming electron beam is focused, but it extends to a region that spreads below it, where electrons scatter and interact with the host material. Cathodoluminescence is potentially generated throughout this volume, the size of which increases dramatically with the electron beam energy. Therefore it should be considered as an extended excitation, although its size can be modulated by a spatially confined generation yield. Most of the CL setups are incorporated in the vacuum chamber of an electron microscope, where the light collector is a parabolic mirror placed on top of the sample. The advantages, challenges and improvement examples of these standard setups are discussed. An overview of applications in cell and molecular biology, geosciences and nanophotonics emphasizes the increasing interest on applying the technique at the nanometer regime. The chapter ends by summarizing the main challenges that cathodoluminescence microscopy encounters for successful imaging of nanostructures on glass, which define the design criteria for our setup. The system details are presented in chapter 2: a brief description of the integrated electron light microscope functionalities and the implementation of the confocal detection path are presented. Explanation of the available acquisition modes, alignment procedures and typical imaging examples serve to establish an operation routine. The effect of the pinhole can be observed by comparing unfiltered and confocal CL images on the same region of a sample. Additionally, the filtering is evaluated without using the electron beam: a laser excitation path included in the setup allows acquiring confocal fluorescence images of a sample with luminescent beads on a glass substrate, for different sizes of the pinhole diameter. Besides efficient CL detection, potential applications of the setup could include: (i) emission localization for excitations with long propagation length, (ii) simultaneous light and electron excitation, (iii) monitoring the effect of electron excitation with subsequent light microscopy, and (iv) the incorporation of light or electron pulses for time-resolved characterization. The use of low energies for the electron excitation probe is proposed in chapter 3 as a strategy to reduce the background CL contribution. This is further investigated with Monte Carlo simulations that show the dependence of the electron interaction volume on the electron beam acceleration voltage. We observe however, that to detect nanostructures with a weak cathodoluminescence signal it is necessary to increase the electron current, which in the low acceleration voltage regime may compromise the spatial resolution. With the low energy approach, individual 30nm phosphor particles are resolved and the high order resonant modes of a gold nanowire on an indium tin oxide (ITO) covered glass microscope slide are detected. For high electron energies, the substrate cathodoluminescence is too strong and overwhelms the signal. Chapter 4 demonstrates confocal filtering as an effective tool for background rejection at high acceleration voltages. The filtering achieved for a given pinhole size is estimated with simulations of the electron interaction volume and measurements of the axial intensity distribution of a phosphor nanoparticle, which acts as a point source. As an illustrative example, a series of CL confocal sections of a gold nanowire on a transparent substrate shows a contrast inversion at the plane where the nanowire is in focus. Here, the highest CL intensity is detected at the position of the structure. The need of a high resolution electron probe is evidenced by acquiring the CL spectral distribution of a gold triangle nano plate, which shows a strong sensitivity to the excitation probe position. Both of the strategies presented in this thesis, the use of low energy excitation and confocal filtering are applicable not only for transparent substrates but for any highly cathodoluminescent material. Chapter 5 explores the use of quantum dots as cathodoluminescent biological markers. In cellular biology, investigation of cellular interactions requires imaging the specific functional proteins on top of the organelles ultrastructure. Therefore, direct correlation between electron and light optical information is a key element for understanding cell function at a molecular level. Among other potential cathodoluminescent markers, quantum dots have the additional advantage that they are already routinely incorporated as bio-labels in fluorescence and consequently, many different bio functionalization possibilities are currently available. Here, we report on the cathodoluminescence detection of bio-functionalized quantum-dots embedded in cells. A high similarity between the fluorescence and cathodoluminescence signals is observed, but the cathodoluminescence signal originates from a smaller sample volume defined by the electron penetration depth. We observe a bleaching of the quantum dots emission under high electron irradiation dose, which so far prevents high magnification imaging. However, recording the fluorescence emission after incremental low dose electron irradiation reveals a complicated dependence of the emission intensity on electron dose, featuring even a regime wherein intensity slightly increases. The origin of this behavior is discussed as a charging mechanism, building on existing models that are also used to explain photo blinking, -bleaching and -brightening of fluorescence from quantum dots. The results presented support the use of cathodoluminescence as a high resolution imaging technique for optical characterization of biological systems. Finally, the main findings on the cathodoluminescence emitted from ITO-covered glass slides, the substrate through this work, are summarized in chapter 6. A dynamic behavior of the intensity and spectral distribution of the emission is observed. Cathodoluminescence measurements at different electron doses reveal a faster cathodoluminescence bleaching with increasing dose, but also the appearance and growth of a new intensity peak at a different position in the spectra. Secondary electron images of the irradiated areas suggest that deposition may be involved in this process. Additionally, experiments with different thicknesses for the ITO conductive layer point to glass as the main responsible for the background emission in our measurements. The results reinforce the importance of sample pre-exposure and confocal filtering for CL characterization at high electron energies.Imaging PhysicsApplied Science
Downhole depth estimation for automated subsurface navigation
No full textThis thesis proposes and evaluates a concept for downhole depth estimation by matching subsurface measurements from two sensors in a bottom hole assembly. The application of a downhole depth estimate in automated subsurface navigation has also been demonstrated. One of the key hurdles in achieving real time subsurface navigation lies in communication bottom neck between surface and downhole. Modern measurement while drilling and logging while drilling tools measure all vital information downhole except depth. Real time availability of depth estimate in the downhole can open new doors for real time automated bit steering and optimization in well drilling operations. The downhole depth estimation concept introduced in this thesis is based on correlation of gamma ray responses from two sensors to estimate average rate of penetration at the bit. This average rate of penetration of the bit is used to estimate distance travelled by the bit. An algorithm is proposed for correlation of gamma ray sensor response. Various parameters in the algorithm are investigated and discussed in detail for optimized performance. The error in the estimation is a result of difference between average and instantaneous rate of penetration as well as wrong correlation. Synthetic sensor response is created from a gamma ray data set to evaluate the algorithm for different noise levels and count rates. The error in estimated depth due to difference between average and instantaneous rate of penetration is approximately 2.2%. The total error is observed to be less than 4% for lower statistical noise levels. To reduce the error associated with proposed system, a nuclear marker - detector system is proposed and evaluated using Monte Carlo based nuclear simulations. In the last section the application of downhole depth for automated well-plan execution through a rotary steerable system is realised.Petroleum EngineeringGeoscience & EngineeringCivil Engineering and Geoscience
High Speed Electron Microscopy: Engineering of a commercial multi-beam scanning electron microscope with transmission imaging
No full textIn this thesis, the design and engineering considerations for a multi-beam scanning electron microscope (MBSEM) are discussed. This microscope can benefit biological research in various ways. It can give new insights into the inner workings of a multitude of biological systems that were hard to get using previously existing instrumentation. For instance, a higher throughput gives the option to do statistical analysis of multiple samples instead of drawing conclusions from only one. The goal of this thesis was to get from a proof of principle to a final system that can actually be used to do the research. It is divided into 5 chapters showing a step-by-step process of getting to the final system as it is now on the market. Chapter 1 is an introduction to the subject showing the current state of the art with respect to high throughput imaging. Chapter 2 Describes a novel imaging method in scanning electron microscopes. This chapter does not focus on the multi-beam application but shows it in the context of the often-used backscatter imaging. In this method, we place the tissue section directly on top of a thin scintillator screen (thinner than 200 μm) which is coated with a conductive layer. The light signal generated by the electrons transmitted through the sample is collected by a high NA objective lens and the light is imaged onto a photon detector outside of the vacuumchamber. A noise model is created to calculate the signal-to-noise ratio and the contrast-tonoise ratio of this imaging method. It shows that the best images are generated around a landing energy of about 5keV. There are some dependencies on sample thickness, staining level, and light collection efficiency which are also explored. This method does lower the resolution in the image to some extent (by a factor of 2 at low energies and thick sections), which is shown at the end of the chapter. Chapter 3 Goes into the considerations that have to be taken into account when dealing with the imaging method from chapter 2. This chapter is applicable to a single beam SEM as much as anMBSEM. A list of possible light detectors is given from which silicon photomultipliers are selected as the best candidate for the MBSEM. Combined with the light detector, multiple options for a scintillator were discussed, from which YAG:CE is selected. Organic scintillators are discarded due to their bleaching behavior due to electron beam irradiation. The surface of the scintillator and the coating layer are shown to have a large impact on image quality. For this reason, the scintillators are ion-beam polished and coated with a Boron layer. Unexpected behavior in the form of scintillator saturation is observed which is then described by a model and connected to the noise model fromchapter 2. Chapter 4 Gives an analysis of all the hardware requirements for a MBSEM. First a measurement of the crosstalk as a function of landing energy and pitch. It is found that a crosstalk of at least 10 % is to be expected in the system. Next, an overview is given for all the parameters that are related to the stage and the light optics. These are then related to the final throughput of the system. Two imaging strategies are described, in one the beam scans in one direction and the stage in the other. In the other strategy, the beams scan like in a regular SEM and are subsequently descanned in the light-optical system. It is found that with a step and scan approach in combination with planned beamshifts, the maximum throughput that can be achieved is around 420 mpix/s. Chapter 5 Shows results from the final prototype system. Alignments are of great importance in any SEMbut even more so in theMBSEM. Therefore a large part of this chapter is dedicated to describing this alignment. This starts with the electron optical alignment of the source and the beam through the column. The grid of beams has to be optimized to show as little as possible distortions to improve system throughput. The scan and descan have to be aligned to the grid axes and the amplitude has to be precisely correct. The beams have to be perfectly aligned to the detector array. On the processing side, a description of how can be compensated for varying dark and gain levels in the detector array. In the end, a final image is shown, consisting of 400 megapixels. Chapter 6 Describes the valorization of the project and all the challenges and choices that were involved.ImPhys/Hoogenboom grou
De toepassing van biomimetica op de woningbouw in overstromingsgebieden
No full textBiomimetica is de wetenschap waarin elementen uit de natuur worden nagebootst met het doel menselijke problemen op te lossen. Evolutionaire ontwikkelingen hebben eraan bijgedragen dat de natuur bestaat uit voornamelijk geoptimaliseerde vormen, structuren en processen. In dit afstudeeronderzoek is biomimicry aangegrepen om tot het ontwerp voor een vloedbestendige woning te komen. In de eerste fase van dit onderzoek zijn de gedragingen van verschillende elementen in de natuur bij (dreigend) hoogwater vastgesteld. Vervolgens is geanalyseerd of deze natuurlijke gedragingen een interessante bijdrage kunnen leveren aan het ontwerp van een vloedbestendige woning. Eén van de elementen uit de natuur die interesse heeft opgewekt is het bananenblad. Deze wordt belast door luchtstroming in plaats van door waterstroming en is aangepast aan deze situatie.De structuur van het bananenblad is zodanig dat deze inscheurt bij hoge windsnelheden. Het belaste oppervlak wordt hierdoor verkleind, zodat de totale belasting op het blad afneemt. De kans dat de plant de storm overleeft wordt groter doordat vitale onderdelen als de stengel minder snel zullen beschadigen. Geanalyseerd is hoe dit concept vertaald kan worden naar een in de woningbouw toe te passen concept. Door de oppervlakte van een gevel te verkleinen zullen de totaalkrachten op de constructie afnemen. Een bijkomend voordeel is de lagere stromingsweerstand van de woning bij afnemende eveloppervlaktes. Dit is een voorwaarde om in bepaalde gebieden te mogen bouwen, bijvoorbeeld in de uiterwaarden van rivieren. Om de bruikbaarheid van een dergelijk concept aan te tonen is in de laatste fase van het afstudeeronderzoek een praktische toepassing gepresenteerd in de vorm van een ontwerpcase. Het resultaat is een ontwerp voor een vrijstaande woning in de uiterwaard van de Nederrijn bij Arnhem. De ontworpen woning beschikt op de begane grond verdieping over lichtgewicht gevels die omhoog kunnen worden geschoven in geval van dreigende hoge waterstanden. Hierdoor transformeert de woning in een paalwoning. De rivier zal bij hoogwater de onderste verdieping benutten voor de doorstroming van het water, iets waar bij de indeling van de woning rekening mee moet worden gehouden. De voornaamste functies van de verdieping zijn het bieden van parkeergelegenheid en opslagruimte.Design and ConstructionCivil Engineering and Geoscience
Concept and design of a beam blanker with integrated photoconductive switch for ultrafast electron microscopy
No full textWe present a new method to create ultrashort electron pulses by integrating a photoconductive switch with an electrostatic deflector. This paper discusses the feasibility of such a system by analytical and numerical calculations. We argue that ultrafast electron pulses can be achieved for micrometer scale dimensions of the blanker, which are feasible with MEMS-based fabrication technology. According to basic models, the design presented in this paper is capable of generating 100 fs electron pulses with spatial resolutions of less than 10 nm. Our concept for an ultrafast beam blanker (UFB) may provide an attractive alternative to perform ultrafast electron microscopy, as it does not require modification of the microscope nor realignment between DC and pulsed mode of operation. Moreover, only low laser pulse energies are required. Due to its small dimensions the UFB can be inserted in the beam line of a commercial microscope via standard entry ports for blankers or variable apertures. The use of a photoconductive switch ensures minimal jitter between laser and electron pulses.Green Open Access added to TU Delft Institutional Repository ‘You share, we take care!’ – Taverne project https://www.openaccess.nl/en/you-share-we-take-care Otherwise as indicated in the copyright section: the publisher is the copyright holder of this work and the author uses the Dutch legislation to make this work public.ImPhys/Charged Particle OpticsImPhys/Quantitative Imagin
Subnanometer-accuracy optical distance ruler based on fluorescence quenching by transparent conductors
No full textAvailable data: Complex refractive index of Indium Tin Oxide, http://dx.doi.org/10.4121/uuid:59febf27-a532-4ac9-8ec0-29d4195b2c8c Transparent conductive oxides (TCOs), such as the well-known indium-tin oxide, find widespread use in modern (nano)technological applications because of their unique combination of negligible optical absorption and good electric conductivity. We, however, show that despite the near-zero imaginary part of the refractive index that is responsible for the material’s transparency, TCOs drastically quench optical emitters when the emitter is within 10 nm from the TCO. Our results reveal that the pure near-field nature of this dissipation makes for an exquisite short-range optical ruler. Previous quenching-based optical rulers, based on interactions with plasmonic or graphene materials, have allowed measuring distances in the 20–100 nm range. Distances below 20 nm have, however, been hard to assess due to poor photon yields or weak absolute variations. We show that TCO-based rulers close this gap, allowing distance measurements with far-field optics in the 1–10 nm distance range with deep subnanometer sensitivity.ImPhys/Imaging PhysicsApplied Science
High Strength Steel Structures – FEM Validation Experiments
No full textStructural and Building EngineeringStructural EngineeringCivil Engineering and Geoscience
Creation of electron pulses with a laser-triggered micro fabricated electron beam deflector
No full textThis thesis is dedicated to the development of a beam blanker for Ultrafast Electron Microscopy. Ultrafast electron microscopy aims to resolve structural dynamics at the nanometer and (sub) picosecond time scale. In these temporal and spatial scales many important processes in physics, chemistry and biology do occur. Examples of these are the interaction of light with small nano-patterned devices, the propagation is acoustic waves and phonons, the dynamics of melting and crystallization of materials. An example in biology is in photosynthesis, i.e. the dynamics of light harvesting complexes.ImPhys/Microscopy Instrumentation & Technique
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