Approaches to overcome flow cytometry limitations in the analysis of cells from veterinary relevant species

BACKGROUND: Flow cytometry is a powerful tool for the multiparameter analysis of leukocyte subsets on the single cell level. Recent advances have greatly increased the number of fluorochrome-labeled antibodies in flow cytometry. In particular, an increase in available fluorochromes with distinct excitation and emission spectra combined with novel multicolor flow cytometers with several lasers have enhanced the generation of multidimensional expression data for leukocytes and other cell types. However, these advances have mainly benefited the analysis of human or mouse cell samples given the lack of reagents for most animal species. The flow cytometric analysis of important veterinary, agricultural, wildlife, and other animal species is still hampered by several technical limitations, even though animal species other than the mouse can serve as more accurate models of specific human physiology and diseases. RESULTS: Here we present time-tested approaches that our laboratory regularly uses in the multiparameter flow cytometric analysis of ovine leukocytes. The discussed approaches will be applicable to the analysis of cells from most animal species and include direct modification of antibodies by covalent conjugation or Fc-directed labeling (Zenon™ technology), labeled secondary antibodies and other second step reagents, labeled receptor ligands, and antibodies with species cross-reactivity. CONCLUSIONS: Using refined technical approaches, the number of parameters analyzed by flow cytometry per cell sample can be greatly increased, enabling multidimensional analysis of rare samples and giving critical insight into veterinary and other less commonly analyzed species. By maximizing information from each cell sample, multicolor flow cytometry can reduce the required number of animals used in a study

Detection of Weak Circumstellar Gas around the DAZ White Dwarf WD 1124-293: Evidence for the Accretion of Multiple Asteroids

Single metal polluted white dwarfs with no dusty disks are believed to be actively accreting metals from a circumstellar disk of gas caused by the destruction of asteroids perturbed by planetary systems. We report, for the first time, the detection of circumstellar Ca~II gas in absorption around the DAZ WD~1124-293, which lacks an infrared excess. We constrain the gas to $>$7 $R_{\rm WD}$ and $<$32000~AU, and estimate it to be at $\sim$54~R$_{\rm WD}$, well within WD~1124-293's tidal disruption radius. This detection is based on several epochs of spectroscopy around the Ca~II H and K lines ($\lambda$=3968\AA, 3933\AA) with the MIKE spectrograph on the Magellan/Clay Telescope at Las Campanas Observatory. We confirm the circumstellar nature of the gas by observing nearby sightlines and finding no evidence for gas from the local interstellar medium. Through archival data we have measured the equivalent width of the two photospheric Ca lines over a period of 11 years. We see $<$ 5-7\% epoch-to-epoch variation in equivalent widths over this time period, and no evidence for long term trends. The presence of a circumstellar gas implies a near edge-on inclination to the system, thus we place limits to short period transiting planetary companions with R $>$ R$_{\rm \oplus}$ using the WASP survey. The presence of gas in orbit around WD~1124-293 implies that most DAZs could harbor planetary systems. Since 25-30\% of white dwarfs show metal line absorption, the dynamical process for perturbing small bodies must be robust.Comment: 31 pages with 9 figures; accepted to Ap

T Cell Migration from Inflamed Skin to Draining Lymph Nodes Requires Intralymphatic Crawling Supported by ICAM-1/LFA-1 Interactions.

T cells are the most abundant cell type found in afferent lymph, but their migration through lymphatic vessels (LVs) remains poorly understood. Performing intravital microscopy in the murine skin, we imaged T cell migration through afferent LVs in vivo. T cells entered into and actively migrated within lymphatic capillaries but were passively transported in contractile collecting vessels. Intralymphatic T cell number and motility were increased during contact-hypersensitivity-induced inflammation and dependent on ICAM-1/LFA-1 interactions. In vitro, blockade of endothelial cell-expressed ICAM-1 reduced T cell adhesion, crawling, and transmigration across lymphatic endothelium and decreased T cell advancement from capillaries into lymphatic collectors in skin explants. In vivo, T cell migration to draining lymph nodes was significantly reduced upon ICAM-1 or LFA-1 blockade. Our findings indicate that T cell migration through LVs occurs in distinct steps and reveal a key role for ICAM-1/LFA-1 interactions in this process

First High Contrast Imaging Using a Gaussian Aperture Pupil Mask

Placing a pupil mask with a gaussian aperture into the optical train of current telescopes represents a way to attain high contrast imaging that potentially improves contrast by orders of magnitude compared to current techniques. We present here the first observations ever using a gaussian aperture pupil mask (GAPM) on the Penn State near-IR Imager and Spectrograph (PIRIS) at the Mt. Wilson 100$^{\prime\prime}$ telescope. Two nearby stars were observed, $\epsilon$ Eridani and $\mu$ Her A. A faint companion was detected around $\mu$ Her A, confirming it as a proper motion companion. Furthermore, the observed H and K magnitudes of the companion were used to constrain its nature. No companions or faint structure were observed for $\epsilon$ Eridani. We found that our observations with the GAPM achieved contrast levels similar to our coronographic images, without blocking light from the central star. The mask's performance also nearly reached sensitivities reported for other ground based adaptive optics coronographs and deep HST images, but did not reach theoretically predicted contrast levels. We outline ways that could improve the performance of the GAPM by an order of magnitude or more.Comment: 8 pages, 4 figures, accepted by ApJ letter

On how the optical depth tunes the effects of ISM neutral atom flow on debris disks

The flux of ISM neutral atoms surrounding stars and their environment affects the motion of dust particles in debris disks, causing a significant dynamical evolution. Large values of eccentricity and inclination can be excited and strong correlations settle in among the orbital angles. This dynamical behaviour, in particular for bound dust grains, can potentially cause significant asymmetries in dusty disks around solar type stars which might be detected by observations. However, the amount of orbital changes due to this non--gravitational perturbation is strongly limited by the collisional lifetime of dust particles. We show that for large values of the disk's optical depth the influence of ISM flow on the disk shape is almost negligible because the grains are collisionally destroyed before they can accumulate enough orbital changes due to the ISM perturbations. On the other hand, for values smaller than $10^{-3}$, peculiar asymmetric patterns appear in the density profile of the disk when we consider 1-10 mum grains, just above the blow-out threshold. The extent and relevance of these asymmetries grow for lower values of the optical depth. An additional sink mechanism, which may prevent the formation of large clumps and warping in the disks is related to the fast inward migration due to the drag component of the forces. When a significant eccentricity is pumped up by the ISM perturbations, the drag forces (Poynting-Robertson and in particular ISM drag) drive the disk particles on fast migrating tracks leading them into the star on a short timescale. It is then expected that disks with small optical depth expand inside the parent body ring all the way towards the star while disks with large optical depth would not significantly extend inside.Comment: accepted for publication in MNRA

The Discovery of the Most Metal-Rich White Dwarf: Composition of a Tidally Disrupted Extrasolar Dwarf Planet

Cool white dwarf stars are usually found to have an outer atmosphere that is practically pure in hydrogen or helium. However, a small fraction have traces of heavy elements that must originate from the accretion of extrinsic material, most probably circumstellar matter. Upon examining thousands of Sloan Digital Sky Survey spectra, we discovered that the helium-atmosphere white dwarf SDSS J073842.56+183509.6 shows the most severe metal pollution ever seen in the outermost layers of such stars. We present here a quantitative analysis of this exciting star by combining high S/N follow-up spectroscopic and photometric observations with model atmospheres and evolutionary models. We determine the global structural properties of our target star, as well as the abundances of the most significant pollutants in its atmosphere, i.e., H, O, Na, Mg, Si, Ca, and Fe. The relative abundances of these elements imply that the source of the accreted material has a composition similar to that of Bulk Earth. We also report the signature of a circumstellar disk revealed through a large infrared excess in JHK photometry. Combined with our inferred estimate of the mass of the accreted material, this strongly suggests that we are witnessing the remains of a tidally disrupted extrasolar body that was as large as Ceres.Comment: 7 pages in emulateapj, 5 figures, accepted for publication in Ap

Analysis of hydrogen-rich magnetic white dwarfs detected in the Sloan Digital Sky Survey

We model the structure of the surface magnetic fields of the hydrogen-rich white dwarfs in the SDSS. We have calculated a grid of state-of-the-art theoretical optical spectra of hydrogen-rich magnetic white dwarfs with magnetic field strengths between 1 MG and 1200 MG for different angles, and for effective temperatures between 7000 K and 50000 K. We used a least-squares minimization scheme with an evolutionary algorithm in order to find the magnetic field geometry best fitting the observed data. We used simple centered dipoles or dipoles which were shifted along the dipole axis to model the coadded SDSS fiber spectrum of each object. We have analysed the spectra of all known magnetic DAs from the SDSS (97 previously published plus 44 newly discovered) and also investigated the statistical properties of magnetic field geometries of this sample. The total number of known magnetic white dwarfs already more than tripled by the SDSS and more objects are expected from a more systematic search. The magnetic fields span a range between ~1 and 900 MG. Our results further support the claim that Ap/Bp population is insufficient in generating the numbers and field strength distributions of the observed MWDs, and either another source of progenitor types or binary evolution is needed. Moreover clear indications for non-centered dipoles exist in about ~50% of the objects which is consistent with the magnetic field distribution observed in Ap/Bp stars.Comment: 15 pages, accepted for publication in A&A. For online version with full appendix figures, see http://www.ari.uni-heidelberg.de/mitarbeiter/bkulebi/papers/12570_online.pd

Dissecting the Moth: Discovery of an off-centered ring in the HD 61005 debris disk with high-resolution imaging

The debris disk known as "The Moth" is named after its unusually asymmetric surface brightness distribution. It is located around the ~90 Myr old G8V star HD 61005 at 34.5 pc and has previously been imaged by the HST at 1.1 and 0.6 microns. Polarimetric observations suggested that the circumstellar material consists of two distinct components, a nearly edge-on disk or ring, and a swept-back feature, the result of interaction with the interstellar medium. We resolve both components at unprecedented resolution with VLT/NACO H-band imaging. Using optimized angular differential imaging techniques to remove the light of the star, we reveal the disk component as a distinct narrow ring at inclination i=84.3 \pm 1.0{\deg}. We determine a semi-major axis of a=61.25 \pm 0.85 AU and an eccentricity of e=0.045 \pm 0.015, assuming that periastron is located along the apparent disk major axis. Therefore, the ring center is offset from the star by at least 2.75 \pm 0.85 AU. The offset, together with a relatively steep inner rim, could indicate a planetary companion that perturbs the remnant planetesimal belt. From our imaging data we set upper mass limits for companions that exclude any object above the deuterium-burning limit for separations down to 0.3". The ring shows a strong brightness asymmetry along both the major and minor axis. A brighter front side could indicate forward-scattering grains, while the brightness difference between the NE and SW components can be only partly explained by the ring center offset, suggesting additional density enhancements on one side of the ring. The swept-back component appears as two streamers originating near the NE and SW edges of the debris ring.Comment: 6 pages, 6 figures. Accepted to Astronomy and Astrophysics letter