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Sum rules and static local-field corrections of electron liquids in two and three dimensions

The dielectric functions of electron liquids which take into account short-range electron-electron correlations via the static local-field corrections are examined in the light of the frequency-moment sum rules. The formation is given for degenerate as well as classical electron liquids in arbitrary (d) spatial dimensions, which is suitable for comparison between the two- and three-dimensional cases. By using the virial equations of state it is shown that such dielectric functions cannot satisfy the compressibility sum rule and the third-frequency-moment sum rule simultaneously. In the degenerate case, the plasmon, single-pair, and multipair contributions to the sum rules are analyzed, and the reason for this incompatibility is discussed.

Iwamoto, N.

Regional Rates of Young US Forest Growth Estimated From Annual Landsat Disturbance History and IKONOS Stereo Imagery

Forests of the Contiguous United States (CONUS) have been found to be a large contributor to the global atmospheric carbon sink. The magnitude and nature of this sink is still uncertain and recent studies have sought to define the dynamics that control its strength and longevity. The Landsat series of satellites has been a vital resource to understand the long-term changes in land cover that can impact ecosystem function and terrestrial carbonstock. We combine annual Landsat forest disturbance history from 1985 to 2011 with single date IKONOS stereoimagery to estimate the change in young forest canopy height and above ground live dry biomass accumulation for selected sites in the CONUS. Our approach follows an approximately linear growth rate following clearing over short intervals and does not estimate the distinct non-linear growth rate over longer intervals.We produced canopy height models by differencing digital surface models estimated from IKONOS stereo pairs with national elevation data (NED). Correlations between height and biomass were established independently using airborne LiDAR, and then applied to the IKONOS-estimated canopy height models. Graphing current biomass against time since disturbance provided biomass accumulation rates. For 20 study sites distributed across five regions of the CONUS, 19 showed statistically significant recovery trends (p is less than 0.001) with canopy growth from 0.26 m yr-1to 0.73 m yr-1. Above ground live dry biomass (AGB) density accumulation ranged from 1.31 t/ha yr-1 to 12.47 t/ha yr-1. Mean forest AGB accumulationwas 6.31 t/ha yr-1 among all sites with significant growth trends. We evaluated the accuracy of our estimates by comparing to field estimated site index curves of growth, airborne LiDAR data, and independent model predictions of C accumulation. Growth estimates found with this approach are consistent with site index curves and total biomass estimates fall within the range of field estimates. This is aviable approach to estimate forest biomass accumulation in regions with clear-cut harvest disturbances.

Neigh, Christopher S. R.

A simulation survey of galaxy interactions

Many carefully selected samples of interacting galaxies have been observed extensively in attempts to clarify whether interaction produces activity in galaxies. Because the sample members represent a wide range of encounter parameters and times, one can then study whether there are correlations between observable encounter features and, for example, Seyfert activity. On the other hand, in theoretical studies, simulations typically deal with either time-consuming detailed modelling of single galaxy pairs or tracing a few model encounters over time. The authors extend the observational survey approach by combining it with a simulation survey. The authors are conducting a survey of model encounters, covering the most important encounter parameters over a wide range. Some parameters, such as companion structure and initial velocity, are demonstratably less important and can be ignored in a first pass. The parameter range must be richly enough sampled so that the authors can evaluate the uniqueness of the observable morphology and velocity structure of the resulting simulated pairs to diagnose unobservable companion orbit parameters. They are using a self-gravitating polar n-body code run on the Cray X-MP at the Alabama Supercomputer Network. For each simulation, the authors have stellar and gas distributions predicted over, typically, a billion years, along with information on gas motions within the disk and any material captured by the companion or lost to the system. Features of disturbed spiral galaxies are sensitive enough to time and encounter parameters so that a match of the simulation survey results to observations can be applied as starting points to infer unobservable orbital or system parameters in actual sample members. This should enable them to examine whether interesting observed properties (Seyfert activity, nuclear star-formation rate) are functions of unobservable dynamical properties which characterize each encounter. Any correlations (or lack of some expected ones) will provide strong clues as to how or whether these phenomena are related to interactions. Aside from its use with such observed samples, this survey should greatly speed determination of initial orbital parameters for more detailed subsequent simulations of individual systems.

Byrd, Gene G.

The high-energy diffuse cosmic gamma-ray background radiation from blazars

We predict the spectrum and flux of the high-energy diffuse cosmic gamma-ray background radiation produced by blazars. Our calculations are based on the Compton Observatory EGRET observations of high-energy gamma-rays from these objects, the Whipple observation of TeV gamma-rays from Mrk 421, and radio population studies of flat-spectrum radio sources. Our statistical analysis gives a result consistent with a linear correlation. We then use the radio luminosity function and redshift evolution to calculate the gamma-ray background. We find that blazars cannot account for the flux level or steep spectrum of the background observed by SAS 2 in the sub-GeV range, but could likely provide the dominant background component in the multi-GeV energy range and above. We predict a differential photon spectral index for this component of about 2 up to about 10 GeV. Above that energy, the spectrum should gradually steepen owing to pair production by interactions of the gamma-rays with intergalactic infrared photons. Using the calculations of this effect by Stecker et al. (1992), we estimate a spectral index of about 3.5 for energies above a TeV.

Stecker, F. W.

Reliability of the Danish Aerospace Corporation Portable Pulmonary Function System

Metabolic gas analysis is a critical component of investigations that measure cardio-pulmonary exercise responses during and after long-duration spaceflight. The primary purpose of the current study was to determine the reliability and intra-subject repeatability of a metabolic gas analysis device, the Portable Pulmonary Function System (PPFS), designed for use on the International Space Station (ISS). The second objective of this study was to directly compare PPFS measurements of expired oxygen and carbon dioxide (FEO2 and FECO2) to values obtained from a well-validated clinical metabolic gas analysis system (ParvoMedics TrueOne (c) [PM]). Eight subjects performed four peak cycle tests to maximal exertion. The first test was used to prescribe work rates for the subsequent test sessions. Metabolic gas analysis for this test was performed by the PM, but samples of FEO2 and FECO2 also were simultaneously collected for analysis by the PPFS. Subjects then performed three additional peak cycle tests, consisting of three 5-min stages designed to elicit 25%, 50%, and 75% maximal oxygen consumption (VO2max) followed by stepwise increases of 25 W/min until subjects reached volitional exhaustion. Metabolic gas analysis was performed using the PPFS for these tests. Intraclass correlation coefficients (ICC), within-subject standard deviations (WS SD), and coefficients of variation (CV%) were calculated for the repeated exercise tests. Mixed model regression analysis was used to compare paired FEO2 and FECO2 values obtained from the PPFS and the PM during the initial test. The ICC values for oxygen consumption (VO2), carbon dioxide production (VCO2), and ventilation (VE) indicate that the PPFS is highly reliable (0.79 to 0.99) for all exercise levels tested; however, ICCs for respiratory exchange ratio (RER) were low ( 0.11 - 0.51), indicating poor agreement between trials during submaximal and maximal exercise. Overall, CVs ranged from 1.6% to 6.7% for all measurements, a finding consistent with reported values that were obtained using other metabolic gas analysis techniques. The PPFS and PM produced comparable FEO2 data; however, there was less agreement between measures of FECO2 obtained from the two devices, particularly at lower CO2 concentrations. The PPFS appears, in practically all respects, to yield highly reliable metabolic gas analysis data. Lower reliability of RER measurements reported in the literature and likely is not a function of the PPFS device. Further examination of PPFS CO2 data is warranted to better understand the limitations of these PPFS measurements. Overall, the PPFS when used for repeated measures of cardio-pulmonary exercise should provide accurate and reliable data for studies of human adaptation to spaceflight.

Portable Pulmonary Function System (PPFS)

Study of Pairwise Deconfliction Metrics to Analyze Air Traffic Complexity in Upper Class E Airspace

Upper Class E Traffic Management (ETM) is envisioned to cooperatively facilitate operations of a diverse set of aerial vehicles, such as high-altitude long-endurance fixed-wing unmanned aircraft (low-speed and high-speed), high-altitude platforms, airships, stratospheric balloons, supersonic unmanned and commercial aircraft, etc., with a wide variety of mission types, performance characteristics, communication, navigation and surveillance capabilities, maneuverability, and on-board avionics in the National Airspace System (NAS) ’above’ 60,000 feet above mean sea level, without an active and direct control from human air traffic controllers. A diverse mixture of aerial vehicle types creates significant challenges in understanding air traffic complexity, which may not correlate strongly with air traffic density. One key step for determining air traffic complexity in upper class E airspace is to first understand pairwise deconfliction metrics such as reachability, reserve area, and reserve flight time for each pair of unique aerial vehicle types under potential conflict. Therefore, pairwise deconfliction metrics are first defined, and analytical equations are derived for conflict resolution using the heading change maneuver. Next, case studies are performed to analyze deconfliction metrics to avoid secondary conflicts in upper class E airspace. The study shows that pairwise deconfliction metrics are functions of maneuverability, performance characteristics, uncertainty in position and velocity, heading angle change, and conflict angle of aerial vehicles. The next step for this research is to build a mathematical model for air traffic complexity using pairwise deconfliction metrics and validate it in an upper Class E simulation environment.

Airspace Complexity

Study of Pairwise Deconfliction Metrics to Analyze Air Traffic Complexity in Upper Class E Airspace

Upper Class E Traffic Management (ETM) is envisioned to cooperatively facilitate operations of a diverse set of aerial vehicles, such as high-altitude long-endurance fixed-wing unmanned aircraft (low-speed and high-speed), high-altitude platforms, airships, stratospheric balloons, supersonic unmanned and commercial aircraft, etc., with a wide variety of mission types, performance characteristics, communication, navigation and surveillance capabilities, maneuverability, and on-board avionics in the National Airspace System (NAS) ’above’ 60,000 feet above mean sea level, without an active and direct control from human air traffic controllers. A diverse mixture of aerial vehicle types creates significant challenges in understanding air traffic complexity, which may not correlate strongly with air traffic density. One key step for determining air traffic complexity in upper class E airspace is to first understand pairwise deconfliction metrics such as reachability, reserve area, and reserve flight time for each pair of unique aerial vehicle types under potential conflict. Therefore, pairwise deconfliction metrics are first defined, and analytical equations are derived for conflict resolution using the heading change maneuver. Next, case studies are performed to analyze deconfliction metrics to avoid secondary conflicts in upper class E airspace. The study shows that pairwise deconfliction metrics are functions of maneuverability, performance characteristics, uncertainty in position and velocity, heading angle change, and conflict angle of aerial vehicles. The next step for this research is to build a mathematical model for air traffic complexity using pairwise deconfliction metrics and validate it in an upper Class E simulation environment.

Airspace Complexity

TACT: A Set of MSC/PATRAN- and MSC/NASTRAN- based Modal Correlation Tools

This paper describes the functionality and demonstrates the utility of the Test Analysis Correlation Tools (TACT), a suite of MSC/PATRAN Command Language (PCL) tools which automate the process of correlating finite element models to modal survey test data. The initial release of TACT provides a basic yet complete set of tools for performing correlation totally inside the PATRAN/NASTRAN environment. Features include a step-by-step menu structure, pre-test accelerometer set evaluation and selection, analysis and test result export/import in Universal File Format, calculation of frequency percent difference and cross-orthogonality correlation results using NASTRAN, creation and manipulation of mode pairs, and five different ways of viewing synchronized animations of analysis and test modal results. For the PATRAN-based analyst, TACT eliminates the repetitive, time-consuming and error-prone steps associated with transferring finite element data to a third-party modal correlation package, which allows the analyst to spend more time on the more challenging task of model updating. The usefulness of this software is presented using a case history, the correlation for a NASA Langley Research Center (LaRC) low aspect ratio research wind tunnel model. To demonstrate the improvements that TACT offers the MSC/PATRAN- and MSC/DIASTRAN- based structural analysis community, a comparison of the modal correlation process using TACT within PATRAN versus external third-party modal correlation packages is presented.

Marlowe, Jill M.

Breadboard Signal Processor for Arraying DSN Antennas

A recently developed breadboard version of an advanced signal processor for arraying many antennas in NASA s Deep Space Network (DSN) can accept inputs in a 500-MHz-wide frequency band from six antennas. The next breadboard version is expected to accept inputs from 16 antennas, and a following developed version is expected to be designed according to an architecture that will be scalable to accept inputs from as many as 400 antennas. These and similar signal processors could also be used for combining multiple wide-band signals in non-DSN applications, including very-long-baseline interferometry and telecommunications. This signal processor performs functions of a wide-band FX correlator and a beam-forming signal combiner. [The term "FX" signifies that the digital samples of two given signals are fast Fourier transformed (F), then the fast Fourier transforms of the two signals are multiplied (X) prior to accumulation.] In this processor, the signals from the various antennas are broken up into channels in the frequency domain (see figure). In each frequency channel, the data from each antenna are correlated against the data from each other antenna; this is done for all antenna baselines (that is, for all antenna pairs). The results of the correlations are used to obtain calibration data to align the antenna signals in both phase and delay. Data from the various antenna frequency channels are also combined and calibration corrections are applied. The frequency-domain data thus combined are then synthesized back to the time domain for passing on to a telemetry receiver

Jongeling, Andre

Simulations of Li+ in Ionic Liquids: Structure, Transport, and Electrochemical Windows

Ionic liquids have been proposed as candidate electrolytes for a number of electrochemical applications. The Li+ solvation structure in these liquids is of central importance to electrolyte properties, like ionic conductivity and electrochemical stability. To this point, we employ simulations at three different size scales to better understand various aspects of the interplay between Li+ solvation structure and dynamics. The smallest systems are Li(Anion)n clusters that are treated with high-accuracy density functional theory (DFT) techniques to provide insight into solvation shell structure through energetics and comparisons to experimental IRRaman spectra. Mid-range sized liquid-phase systems (12-24 ion pairs) are treated with DFT molecular dynamics (MD) to provide temperature-dependent insight into Li+ solvation structure, diffusion, and electrochemical window. The largest systems (144-216 ion pairs) are treated with polarizable MD simulations to evaluate the influence of Li-networks on structure and provide size independent values of transport properties. We perform this procedure on three technologically important ionic liquids and comment on property correlations with solvation structure.

batteries

The binding energies of Cu(+)-(H2O)n and Cu(+)-(NH3)n (n = 1-4)

The successive binding energies of up to four H2O and NH3 ligands to Cu(+) are computed at the self-consistent-field and modified coupled-pair functional levels. The most stable structures are those where all ligands are equivalent. Replacing Cu(+) by a point charge gives binding energies that are in good agreement with ab initio and experimental results, and is consistent with bonding that is largely charge dipole in nature. About two-thirds of the large reduction in ligand binding energy between the second and third ligand is due to ligand-ligand repulsion, while one-third is due to increased metal-ligand repulsion resulting from a loss of sd-sigma hybridization. The first and second ligand binding energies increase substantially at the correlated level due to an improved description of sd-sigma hybridization.

Bauschlicher, Charles W., Jr.

Optimal Time Advance In Terminal Area Arrivals: Throughput vs. Fuel Savings

The current operational practice in scheduling air traffic arriving at an airport is to adjust flight schedules by delay, i.e. a postponement of an aircrafts arrival at a scheduled location, to manage safely the FAA-mandated separation constraints between aircraft. To meet the observed and forecast growth in traffic demand, however, the practice of time advance (speeding up an aircraft toward a scheduled location) is envisioned for future operations as a practice additional to delay. Time advance has two potential advantages. The first is the capability to minimize, or at least reduce, the excess separation (the distances between pairs of aircraft immediately in-trail) and thereby to increase the throughput of the arriving traffic. The second is to reduce the total traffic delay when the traffic sample is below saturation density. A cost associated with time advance is the fuel expenditure required by an aircraft to speed up. We present an optimal control model of air traffic arriving in a terminal area and solve it using the Pontryagin Maximum Principle. The admissible controls allow time advance, as well as delay, some of the way. The cost function reflects the trade-off between minimizing two competing objectives: excess separation (negatively correlated with throughput) and fuel burn. A number of instances are solved using three different methods, to demonstrate consistency of solutions.

Sadovsky, Alexander V .

Long-Term Validation and Variability of the Shortwave and Longwave Radiation Data of the GEWEX Surface Radiation Budget (SRB) Project

In this investigation, we make systematic Surface Radiation Budget-Baseline Surface Radiation Network (SRB-BSRN), Surface Radiation Data Centre (SRB-WRDC) and Surface Radiation Budget-Global Energy Balance Archive (SRB-GEBA) comparisons for both shortwave and longwave daily and monthly mean radiation fluxes at the Earth's surface. We first have an overview of all the comparable pairs of data in scatter or scatter density plots. Then we show the time series of the SRB data at grids in which there are ground sites where longterm records of data are available for comparison. An overall very good agreement between the SRB data and ground observations is found. To see the variability of the SRB data during the 21.5 years, we computed the global mean and its linear trend. No appreciable trend is detected at the 5% level. The empirical orthogonal functions (EOF) of the SRB deseasonalized shortwave downward flux are computed over the Pacific region, and the first EOF coefficient is found to be correlated with the ENSO Index at a high value of coefficient of 0.7083.

Zhang, Taiping

Support Routines for In Situ Image Processing

This software consists of a set of application programs that support ground-based image processing for in situ missions. These programs represent a collection of utility routines that perform miscellaneous functions in the context of the ground data system. Each one fulfills some specific need as determined via operational experience. The most unique aspect to these programs is that they are integrated into the large, in situ image processing system via the PIG (Planetary Image Geometry) library. They work directly with space in situ data, understanding the appropriate image meta-data fields and updating them properly. The programs themselves are completely multimission; all mission dependencies are handled by PIG. This suite of programs consists of: (1)marscahv: Generates a linearized, epi-polar aligned image given a stereo pair of images. These images are optimized for 1-D stereo correlations, (2) marscheckcm: Compares the camera model in an image label with one derived via kinematics modeling on the ground, (3) marschkovl: Checks the overlaps between a list of images in order to determine which might be stereo pairs. This is useful for non-traditional stereo images like long-baseline or those from an articulating arm camera, (4) marscoordtrans: Translates mosaic coordinates from one form into another, (5) marsdispcompare: Checks a Left Right stereo disparity image against a Right Left disparity image to ensure they are consistent with each other, (6) marsdispwarp: Takes one image of a stereo pair and warps it through a disparity map to create a synthetic opposite- eye image. For example, a right eye image could be transformed to look like it was taken from the left eye via this program, (7) marsfidfinder: Finds fiducial markers in an image by projecting their approximate location and then using correlation to locate the markers to subpixel accuracy. These fiducial markets are small targets attached to the spacecraft surface. This helps verify, or improve, the pointing of in situ cameras, (8) marsinvrange: Inverse of marsrange . given a range file, re-computes an XYZ file that closely matches the original. . marsproj: Projects an XYZ coordinate through the camera model, and reports the line/sample coordinates of the point in the image, (9) marsprojfid: Given the output of marsfidfinder, projects the XYZ locations and compares them to the found locations, creating a report showing the fiducial errors in each image. marsrad: Radiometrically corrects an image, (10) marsrelabel: Updates coordinate system or camera model labels in an image, (11) marstiexyz: Given a stereo pair, allows the user to interactively pick a point in each image and reports the XYZ value corresponding to that pair of locations. marsunmosaic: Extracts a single frame from a mosaic, which will be created such that it could have been an input to the original mosaic. Useful for creating simulated input frames using different camera models than the original mosaic used, and (12) merinverter: Uses an inverse lookup table to convert 8-bit telemetered data to its 12-bit original form. Can be used in other missions despite the name.

Deen, Robert G.

Validation of Cardiovascular Parameters during NASA's Functional Task Test

Microgravity exposure causes physiological deconditioning and impairs crewmember task performance. The Functional Task Test (FTT) is designed to correlate these physiological changes to performance in a series of operationally-relevant tasks. One of these, the Recovery from Fall/Stand Test (RFST), tests both the ability to recover from a prone position and cardiovascular responses to orthostasis. PURPOSE: Three minutes were chosen for the duration of this test, yet it is unknown if this is long enough to induce cardiovascular responses similar to the operational 5 min stand test. The purpose of this study was to determine the validity and reliability of heart rate variability (HRV) analysis of a 3 min stand and to examine the effect of spaceflight on these measures. METHODS: To determine the validity of using 3 vs. 5 min of standing to assess HRV, ECG was collected from 7 healthy subjects who participated in a 6 min RFST. Mean R-R interval (RR) and spectral HRV were measured in minutes 0-3 and 0-5 following the heart rate transient due to standing. Significant differences between the segments were determined by a paired t-test. To determine the reliability of the 3-min stand test, 13 healthy subjects completed 3 trials of the FTT on separate days, including the RFST with a 3 min stand. Analysis of variance (ANOVA) was performed on the HRV measures. One crewmember completed the FTT before a 14-day mission, on landing day (R+0) and one (R+1) day after returning to Earth. RESULTS VALIDITY: HRV measures reflecting autonomic activity were not significantly different during the 0-3 and 0-5 min segments. RELIABILITY: The average coefficient of variation for RR, systolic (SBP) and diastolic blood pressures during the RFST were less than 8% for the 3 sessions. ANOVA results yielded a greater inter-subject variability (p<0.006) than inter-session variability (p>0.05) for HRV in the RFST. SPACEFLIGHT: Lower RR and higher SBP were observed on R+0 in rest and stand. On R+1, both RR and SBP trended towards preflight rest and stand values. Postflight HRV showed higher LF/HF for rest and stand and lower HFnu during rest. CONCLUSION: These studies show that a 3 min stand delivers repeatable HRV data in the context of this larger series of FTT tests. Spaceflight-induced changes in blood pressure, RR and autonomic function (HRV) are evident from the RFST.

Arzeno, N. M.

Interphase Chromosome Conformation and Chromatin-Chromatin Interactions in Human Epithelial Cells Cultured Under Different Gravity Conditions

Microgravity has been shown to alter global gene expression patterns and protein levels both in cultured cells and animal models. It has been suggested that the packaging of chromatin fibers in the interphase nucleus is closely related to genome function, and the changes in transcriptional activity are tightly correlated with changes in chromatin folding. This study explores the changes of chromatin conformation and chromatin-chromatin interactions in the simulated microgravity environment, and investigates their correlation to the expression of genes located at different regions of the chromosome. To investigate the folding of chromatin in interphase under various culture conditions, human epithelial cells, fibroblasts, and lymphocytes were fixed in the G1 phase. Interphase chromosomes were hybridized with a multicolor banding in situ hybridization (mBAND) probe for chromosome 3 which distinguishes six regions of the chromosome as separate colors. After images were captured with a laser scanning confocal microscope, the 3-dimensional structure of interphase chromosome 3 was reconstructed at multi-mega base pair scale. In order to determine the effects of microgravity on chromosome conformation and orientation, measures such as distance between homologous pairs, relative orientation of chromosome arms about a shared midpoint, and orientation of arms within individual chromosomes were all considered as potentially impacted by simulated microgravity conditions. The studies revealed non-random folding of chromatin in interphase, and suggested an association of interphase chromatin folding with radiation-induced chromosome aberration hotspots. Interestingly, the distributions of genes with expression changes over chromosome 3 in cells cultured under microgravity environment are apparently clustered on specific loci and chromosomes. This data provides important insights into how mammalian cells respond to microgravity at molecular level.

International Space Station

On the joint spectral density of bivariate random sequences

For univariate random sequences, the power spectral density acts like a probability density function of the frequencies present in the sequence. This dissertation extends that concept to bivariate random sequences. For this purpose, a function called the joint spectral density is defined that represents a joint probability weighing of the frequency content of pairs of random sequences. Given a pair of random sequences, the joint spectral density is not uniquely determined in the absence of any constraints. Two approaches to constraining the sequences are suggested: (1) assume the sequences are the margins of some stationary random field, (2) assume the sequences conform to a particular model that is linked to the joint spectral density. For both approaches, the properties of the resulting sequences are investigated in some detail, and simulation is used to corroborate theoretical results. It is concluded that under either of these two constraints, the joint spectral density can be computed from the non-stationary cross-correlation.

Aalfs, David D.

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.