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Two-Photon Fluorescence Microscope for Microgravity Research

A two-photon fluorescence microscope has been developed for the study of biophysical phenomena. Two-photon microscopy is a novel form of laser-based scanning microscopy that enables three-dimensional imaging without many of the problems inherent in confocal microscopy. Unlike one-photon optical microscopy, two-photon microscopy utilizes the simultaneous nonlinear absorption of two near-infrared photons. However, the efficiency of two-photon absorption is much lower than that of one-photon absorption, so an ultra-fast pulsed laser source is typically employed. On the other hand, the critical energy threshold for two-photon absorption leads to fluorophore excitation that is intrinsically localized to the focal volume. Consequently, two-photon microscopy enables optical sectioning and confocal performance without the need for a signal-limiting pinhole. In addition, there is a reduction (relative to one-photon optical microscopy) in photon-induced damage because of the longer excitation wavelength. This reduction is especially advantageous for in vivo studies. Relative to confocal microscopy, there is also a reduction in background fluorescence, and, because of a reduction in Rayleigh scattering, there is a 4 increase of penetration depth. The prohibitive cost of a commercial two-photon fluorescence-microscope system, as well as a need for modularity, has led to the construction of a custom-built system (see Figure 1). This system includes a coherent mode-locked titanium: sapphire laser emitting 120-fs-duration pulses at a repetition rate of 80 MHz. The pulsed laser has an average output power of 800 mW and a wavelength tuning range of 700 to 980 nm, enabling the excitation of a variety of targeted fluorophores. The output from the laser is attenuated, spatially filtered, and then directed into a confocal scanning head that has been modified to provide for side entry of the laser beam. The laser output coupler has been replaced with a dichroic filter that reflects the longer-wavelength excitation light and passes the shorter-wavelength fluorescence light. Also, the confocal pinhole has been removed to increase the signal strength. The laser beam is scanned by a twoperpendicular- axis pair of galvanometer mirrors through a pupil transfer lens into the side port of an inverted microscope. Finally, the beam is focused by a 63-magnification, 1.3-numerical- aperture oil-immersion objective lens onto a specimen. The pupil transfer lens serves to match the intermediate image planes of the scanning head and the microscope, and its location is critical. In order to maximize the quality of the image, (that is, the point spread function of the objective lens for all scan positions), the entire system was modeled in optical-design software, and the various free design parameters (the parameters of the spatial-filter components as well as the separations of all of the system components) were determined through an iterative optimization process. A modular design was chosen to facilitate access to the optical train for future fluorescence correlation spectroscopy and fluorescence-lifetime experiments.

Fischer, David G.↗

Quantitative Species Measurements in Microgravity Combustion Flames using Near-Infrared Diode Lasers

Understanding the physical phenomena controlling the ignition and spread of flames in microgravity has importance for space safety as well as for characterizing dynamical and chemical combustion processes which are normally masked by buoyancy and other gravity-related effects. Unfortunately, combustion is highly complicated by fluid mechanical and chemical kinetic processes, requiring the use of numerical modeling to compare with carefully designed experiments. More sophisticated diagnostic methods are needed to provide the kind of quantitative data necessary to characterize the properties of microgravity combustion as well as provide accurate feedback to improve the predictive capabilities of the models. Diode lasers are a natural choice for use under the severe conditions of low gravity experiments. Reliable, simple solid state operation at low power satisfies the operational restrictions imposed by drop towers, aircraft and space-based studies. Modulation wavelength absorption spectroscopy (WMS) provides a means to make highly sensitive and quantitative measurements of local gas concentration and, in certain cases, temperature. With near-infrared diode lasers, detection of virtually all major combustion species with extremely rapid response time is possible in an inexpensive package. Advancements in near-infrared diode laser fabrication technology and concurrent development of optical fibers for these lasers led to their use in drop towers. Since near-infrared absorption line strengths for overtone and combination vibrational transitions are weaker than the mid-infrared fundamental bands, WMS techniques are applied to increase detection sensitivity and allow measurement of the major combustion gases. In the first microgravity species measurement, Silver et al. mounted a fiber-coupled laser at the top of the NASA 2.2-sec drop tower and piped the light through a single-mode fiber to the drop rig. A fiber splitter divided the light into eight channels that directed the laser beam across a methane or propane diffusion jet flame. The light beams were recaptured by a set of gradient index lenses, coupled back into separate fiber optic lines, and transmitted back to detectors and electronics in the instrument package. In these experiments a 6-mm od fiber cable (containing the nine optical fibers) fell with the drop rig. Using separate detection and demodulation channels, spatial and temporal (up to 20 Hz) maps of water vapor and methane concentrations were obtained at differing heights in the flames. While this apparatus was useful from a demonstration standpoint, several drawbacks needed attention before useful scientific measurements could be obtained. First, eight lines of sight are somewhat insufficient for detailing the spatial profiles of the gas. Second, multiple detection channels operating in parallel are both expensive and present a challenge for accurate calibration. As a result, a newer scanning system was developed in our first contract under this program. The primary characteristic of this system is that it contains a single detection channel and achieves "continuous" spatial resolution by scanning the laser beam across the flame region, then directing this beam onto a single detector. Thus spatial measurements are converted to a temporal series of data. The true spatial resolution is limited only by the beam diameter and width of the sweep. In these experiments the beam is focused to about 1-mm diameter and scans across a region up to 4-cm wide.

Silver, Joel A.↗

Chapter 10 - Remote Sensing Measurements of Aerosol Properties

Satellite instruments have proven especially capable at monitoring the quantity of airborne particles in columns of atmosphere, globally. This chapter describes the principles of satellite measurements and retrieval algorithms, and surveys current instruments and their capabilities. We outline the issues associated with retrieval algorithms, such as surface characterization and aerosol proximity to clouds, and the challenges with interpretation of the results. The relationship between measured aerosol properties and climate-relevant aerosol properties simulated in models is outlined, as well as how measurements are used to evaluate models. Most space-based aerosol instruments are passive sensors that measure reflected sunlight at multiple wavelengths, some at multiple viewing angles. A few are active sensors that send out their own laser light and measure the returned signal. Except when clouds are present, the excess amount of light scattered back to space, beyond that expected from the surface and atmospheric gas, is attributed to aerosol. Satellite measurements are used in many ways in aerosol research. They often provide the only method for monitoring hazardous phenomena such as major wildfire and volcanic eruption plumes, especially in remote areas. Stable, long-term, near-global-scale satellite data records make it possible to identify regional and global aerosol trends. Aerosol radiative effects on climate can be quantified on a near-global scale and used to estimate the strength of aerosol–radiation and aerosol–cloud interactions as well as to evaluate climate model simulations of these interactions. Aerosol-type mapping from satellite imagery is helpful for source attribution, model validation, and to constrain particle light-absorption properties that are essential for radiative forcing calculations. The range of aerosol properties retrieved from satellite observations has grown considerably since the first global estimates of aerosol optical depth (τ a) over ocean were made in the late 1970s. Methods for retrieving particle size and light-absorption properties were explored in the 1990s using multispectral, multi-angle observations, and polarization in visible and near-infrared wavelengths. Sensitivity to particle light absorption, primarily from black or brown carbon content, improved with the inclusion of UV channels, and sensitivity to very thin aerosol layers in the upper troposphere and lower stratosphere was advanced with the use of limb-sounding instruments and active sensors. There are limitations to every measurement technique, including satellite aerosol remote sensing. For wide-swath, passive instruments, aerosol retrievals near clouds can present substantial challenges as far as 15 km away due to cloud-scattered light contaminating the signal. In nearly all cases, retrievals over bright snow and ice surfaces are precluded because surface reflectance uncertainties can overwhelm the aerosol signal. Similarly, meteorological cloud is identified and masked out where possible. Data from passive sensors also lack vertical resolution except those that view toward the limb or where multi-angle imagery is acquired over plumes from wildfires, erupting volcanoes, and wind-blown dust. Yet, passive sensors provide vastly more coverage than the active instruments that mitigate these issues. Particle microphysical information is qualitative from all remote sensing techniques, relying on proxies to infer particle composition, hygroscopicity, and the amount of light-absorbing material. Further, particles smaller than about 200 nm diameter cannot be distinguished from atmospheric gas molecules with remote sensing, which hinders studies of cloud condensation nuclei and their effects on clouds. Most satellite instruments dedicated to aerosol observations are in low-Earth, near-polar, sun-synchronous orbits, which means they cross the equator at the same local time each day. Most are set on cycles that repeat approximately every 16 days, which makes it difficult to monitor aerosol evolution locally. Geostationary satellites make it possible to observe changes occurring from minutes to hours over regions up to 8000 km in size, but lack coverage of high latitudes, and often provide more limited constraints on aerosol properties. Ground-truth data are vital for satellite aerosol-retrieval validation. The AErosol RObotic NETwork (AERONET) of sun photometers was created in 1993 and has become an established global network of over 350 instruments for validating satellite measurements. The network, as well as global networks of ground-based lidars, solar flux radiometers and other sun photometers, are widely used for evaluating global satellite retrievals and model simulations. NASA's Earth Observing System (EOS) program beginning in 1999 led to improvements in reliability, spatial resolution, and spectral resolution (and hence, to improved particle size discrimination and light absorption properties). Satellite payloads include advanced broad-swath and multi-angle imagers, along with the first space-based active sensor focused largely on long-term aerosol monitoring. Since about 2002, Europe's SENTINEL and operational meteorological satellite fleets are also providing sustained aerosol observations, with planned continuation until at least 2030. Satellite remote sensing instruments offer valuable data for evaluating aerosol representations in global climate models. They have been used to assess aerosol optical and physical properties, trends and distributions, and are applied increasingly as direct model constraints in data assimilation to create global aerosol reanalysis products. Aerosol optical depth is the most common quantity adopted for routine model evaluation, including multiwavelength data to loosely constrain particle-size distributions. These evaluations of multiple models have revealed general biases in their regional aerosol amounts and seasonal patterns of transport and removal. Although satellite measurements have near-global coverage, substantial errors can be introduced into the model observation comparison unless attention is paid to spatial and temporal collocation, cloud screening, subgrid-scale variability, and measurement uncertainties that vary with retrieval conditions.

aerosol properties↗

Shockwave/Boundary-Layer Interaction Studies Performed in the NASA Langley 20-Inch Mach 6 Air Tunnel

This paper highlights results from a collaborative study performed by The University of Tennessee Space Institute (UTSI) and NASA Langley Research Center on the Shockwave/Boundary-Layer Interaction (SWBLI) generated by a cylindrical protuberance on a flat plate in a Mach 6 flow. The study was performed in the 20-Inch Mach 6 Air Tunnel at NASA Langley Research Center and consisted of two separate entries. In the first entry, simultaneous high-speed schlieren and high-speed pressure-sensitive paint (PSP) imaging – which was performed for the first time in the 20-Inch Mach 6 facility at NASA Langley – were performed as well as simultaneous high-speed schlieren and oil-flow imaging. In the second entry, the model configuration was modified to increase the size of the interaction region. High-speed schlieren and infrared thermography (IR) surface imaging were performed in this second entry. The goal of these tests was to characterize the SBLI in the presence of a laminar, transitional, and turbulent boundary layer using high-speed optical imaging techniques. AoA = sting angle-of-attack (°) dcylinder = cylinder diameter (mm) dtrip = cylindrical tripping element diameter (mm) Δshock = shock stand-off distance (mm) hcylinder = cylinder height (mm) htrip = cylindrical tripping element height (mm) HSS = high-speed schlieren M∞ = freestream Mach number PSP = pressure-sensitive paint Re∞ = freestream unit Reynolds number (m-1) SWBLI = shockwave/boundary-layer interaction θplate = model plate angle (°) Introduction his paper highlights two experimental entries performed in the 20-Inch Mach 6 Air Blowdown Tunnel at NASA Langley Research Center in collaboration with The University of Tennessee Space Institute (UTSI). The purpose of these entries was to characterize the dynamic shockwave/boundary-layer interaction (SWBLI) between a vertical cylinder on a flat plate and laminar, transitional (XSWBLI), and turbulent (SWTBLI) boundary layers with a freestream Mach number of 6 using non-intrusive optical diagnostics. Experiments performed by Murphree et al.1,2 were among the first to specifically characterize XSWBLI induced by a vertical cylinder on a flat plate geometry using several optical measurement techniques. Recent optical studies of XSWBLI phenomenon have been performed by UTSI at Mach 2 in their low-enthalpy blow wind tunnel3-8 and by Texas A&M University and UTSI at Mach numbers of 6 and 7 in their Adjustable Contour Expansion wind tunnel.9 The experiments described in this paper were intended to complement previous studies by expanding the freestream unit Reynolds number range, Re∞, over which the XSWBLI phenomena has been observed. Additionally these experiments, made possible under NASA’s new facility funding model under the Aeronautics Evaluation and Test Capabilities (AETC) project, promoted collaboration between university and NASA researchers. The initial entry in the 20-Inch Mach 6 Air Tunnel at NASA Langley occurred in December of 2016. Originally, testing was to occur in November of 2016 in the 31-Inch Mach 10 Air Tunnel at NASA Langley. This facility was chosen so that the XSWBLI phenomenon could be observed at much higher Mach numbers than had previously been attempted in ground test experiments. The model selected for this experiment, a 10° half-angle wedge with a sharp leading edge (described in detail in section II.B), had previously been used by Danehy et al. [10] for boundary layer transition studies using the nitric oxide planar laser-induced fluorescence (NO PLIF) flow visualization technique. In that work, it was determined that transition could be induced downstream of a single htrip = 1-mm tall, dtrip = 4-mm diameter cylindrical tripping element and that the streamwise location of the transition could be changed for a single Re∞ by changing the model angle-of-attack (AoA) (see Fig. A3 in Ref. [10] for more details). Based on the findings of that work, a decision was made to use the wedge model with the cylindrical tripping element to trip the boundary layer flow ahead of a cylindrical protuberance in order to achieve a XSWBLI. Unfortunately, the 31-Inch Mach 10 facility had been taken offline for repairs in October of 2016 and a decision was made to move the test to the 20-Inch Mach 6 facility. Since the behavior of the boundary layer with the chosen model configuration had not been studied before in that facility and the available test time was limited, the entry was considered to be exploratory and was used to collect spatially-resolved and time-resolved flow and surface visualization data that would be used to inform a second entry. Test techniques included simultaneous high-speed schlieren (HSS) captured at 160 kHz and high-speed pressure sensitive paint captured at 10 kHz as well as oil flow visualization, captured at 750 Hz. The second entry in the 20-Inch Mach 6 facility occurred in June and July of 2017. In this follow-on test, modifications to the wind tunnel model were made based on observations made during the first entry and included removing the cylindrical tripping element, increasing the size of the cylinder used to induce the SWBLI to increase the size of the interaction while simultaneously improving spatial resolution, and using a swept ramp array, similar to that described in Ref. [11], to trip the flow to turbulence. Simultaneous HSS (captured at 140 kHz, 100 kHz, and 40 kHz) and conventional IR thermography (captured at 30 Hz) imaging were performed simultaneously in this follow-on entry. This paper is intended to serve as a summary of the work performed during these two entries, to detail lessons learned from each entry, and to highlight some of the datasets acquired. Details on the experimental setup, model configuration, and techniques used are provided. Papers providing a more rigorous analysis of data acquired during the second entry, including statistical, spectral, and modal decomposition methods, can be found in Refs. [12,13]. An entry examining XSWBLI in the 31-Inch Mach 10 Blowdown Wind Tunnel facility is currently planned for mid-to-late calendar year 2019, pending the success of facility repairs. The work performed and described in this paper and the upcoming entry in the 31-Inch Mach 10 facility at NASA Langley have been made possible by NASA’s new facility funding model under the Aeronautics Evaluation and Test Capabilities (AETC) project. Wind Tunnel Facility All experiments discussed in this paper were performed in the 20-Inch Mach 6 Air Tunnel at NASA Langley Research Center. Specific details pertaining to this facility can be found in Refs. [14,15], with only a brief description of the facility provided here. For both entries, the nominal freestream unit Reynolds number was varied between 1.8×106 m-1 (0.5×106 ft-1) and 26.3×106 m-1 (8×106 ft-1). The nominal stagnation pressure was varied between 0.21 MPa and 3.33 MPa and the nominal stagnation temperature was varied between 480 K and 520 K to achieve the desired Re∞ condition. For all runs, the nominal freestream Mach number was 6. The nearly square test section is 520.7-mm (20.5-inches) wide by 508-mm (20-inches) high. Two 431.8-mm (17-inch) diameter windows made of Corning 7940, Grade 5F schlieren-quality glass serve as the side walls of the tunnel and provide optical access for the high-speed schlieren measurements. A rectangular window made of the same material as the side windows served as the top wall of the test section and provided optical access for the high-speed PSP and oil flow measurements. For the second entry, this top window was replaced with a Zinc Selenide (ZnSe) window with an anti-reflection coating capable of passing IR wavelengths between 8μm and 12μm with greater than 98% transmittance. The model was sting supported by a strut attached to a hydraulic system that allows for the model pitch angle to be adjusted between -5° to +55°. For the first entry, an initial pitch/pause sweep of the model AoA was performed to observe the resulting SWBLI. Ultimately, however, the sting pitch angle for this entry was fixed at +10.0° so that the angle of the top surface of the wedge relative to the streamwise axis of the tunnel (referred to herein as the plate angle, θplate), was θplate = 0°. For the second entry, θplate = 0° and θplate = -13.25° were initially tested with the swept ramp array (discussed in the following section) to determine which orientation produced conditions most favorable for XSWBLI to occur based on the heating signatures observed over the top surface of the model in the IR thermography images. Based on these initial tests, θplate = -13.25° was set for the remainder of the runs in the second entry. For both entries, any model changes were performed in a housing located beneath the closed test section. Prior to performing a run of the tunnel, the housing was sealed and the tunnel started. Once the appropriate freestream conditions were achieved, the model was injected into the test section using a hydraulic injection system. B. Model Geometry For all runs, a 10° half-angle (20° full-angle) wedge model with a sharp leading edge was used. The model is described in detail in Refs. [10,16]. The top surface of the sharp leading edge of the model extended 47.8 mm from its upstream-most edge to a junction with the upstream edge of a stainless steel top plate that then extended an (a) (c) (b) Fig. 1 (a) Schematic of top surface of wedge model with gas seeding insert, (b) perspective view of the model in the 20-Inch Mach 6 tunnel with centerline pressure orifices on sharp leading edge, and (c) a perspective view of the model with stainless steel (top) and SLA middle insert (bottom) during the first entry. Flow occurs from left to right.

Bathel, Brett F.↗