Antibody-based detection of protein phosphorylation status to track the efficacy of novel therapies using nanogram protein quantities from stem cells and cell lines

This protocol describes a highly reproducible antibody-based method that provides protein level and phosphorylation status information from nanogram quantities of protein cell lysate. Nanocapillary isoelectric focusing (cIEF) combines with UV-activated linking chemistry to detect changes in phosphorylation status. As an example application, we describe how to detect changes in response to tyrosine kinase inhibitors (TKIs) in the phosphorylation status of the adaptor protein ​CrkL, a major substrate of the oncogenic tyrosine kinase ​BCR-​ABL in chronic myeloid leukemia (CML), using highly enriched CML stem cells and mature cell populations in vitro. This protocol provides a 2.5 pg/nl limit of protein detection (<0.2% of a stem cell sample containing <104 cells). Additional assays are described for phosphorylated tyrosine 207 (pTyr207)-​CrkL and the protein tyrosine phosphatase ​PTPRC/​CD45; these assays were developed using this protocol and applied to CML patient samples. This method is of high throughput, and it can act as a screen for in vitro cancer stem cell response to drugs and novel agents

ISO observations of a sample of Compact Steep Spectrum and GHz Peaked Spectrum Radio Galaxies

We present results from observations obtained with ISOPHOT, on board the ISO satellite, of a representative sample of seventeen CSS/GPS radio galaxies and of a control sample of sixteen extended radio galaxies spanning similar ranges in redshift (0.2 = 10^26 W/Hz). The observations have been performed at lambda = 60, 90, 174 and 200 microns. Seven of the CSS/GPS sources have detections >= 3 sigma at one or more wavelengths, one of which is detected at >= 5 sigma. By co-adding the data we have obtained average flux densities at the four wavelengths. We found no evidence that the FIR luminosities of the CSS/GPS sources are significantly different from those of the extended objects and therefore there is not any support for CSS/GPS sources being objects "frustrated" by an abnormally dense ambient medium. The two samples were then combined, providing FIR information on a new sample of radio galaxies at intermediate redshifts. We compare this information with what previously known from IRAS and discuss the average properties of radio galaxies in the redshift range 0.2 - 0.8. The FIR emission cannot be accounted for by extrapolation of the synchrotron radio spectrum and we attribute it to thermal dust emission. The average FIR luminosity is >= 6*10^11 L_sun. Over the observed frequency range the infrared spectrum can be described by a power law with spectral index alpha >~1.0 +/- 0.2. Assuming the emission to be due to dust, a range of temperatures is required, from >=80 K to \~25 K. The dust masses required to explain the FIR emission range from 5*10^5 M_sun for the hotter component up to 2*10^8 M_sun for the colder one. (abridged)Comment: Astronomy & Astrophysics, in press, 16 pages, 2 Figure

Evidence for Ordered Magnetic Fields in the Quasar Environment

At a distance of 20 pc from the purported supermassive black hole powering quasars, temperatures and densities are inferred from optical observations to be ~10**4 K and ~10**4 cm**-3. Here we present Very Long Baseline Interferometry radio observations revealing organized magnetic fields on the parsec scale in the hot plasma surrounding the quasar OQ172 (1442+101). These magnetic fields rotate the plane of polarization of the radio emission coming from the core and inner jet of the quasar. The derived rotation measure (RM) is 40,000 rad m**-2 in the rest frame of the quasar. Only 10 mas (a projected distance of 68 pc) from the nucleus the jet absolute values of RM fall to less than 100 rad m**-2.Comment: in press at ApJ Letters, 12 page LaTeX document includes 4 postscript figure

Observations of HI Absorbing Gas in Compact Radio Sources at Cosmological Redshifts

We present an overview of the occurrence and properties of atomic gas associated with compact radio sources at redshifts up to z=0.85. Searches for HI 21cm absorption were made with the Westerbork Synthesis Radio Telescope at UHF-high frequencies (725-1200 MHz). Detections were obtained for 19 of the 57 sources with usable spectra (33%). We have found a large range in line depths, from tau=0.16 to tau<=0.001. There is a substantial variety of line profiles, including Gaussians of less than 10km/s, to more typically 150km/s, as well as irregular and multi-peaked absorption profiles, sometimes spanning several hundred km/s. Assuming uniform coverage of the entire radio source, we obtain column depths of atomic gas between 1e19 and 3.3e21(Tsp/100K)(1/f)cm^(-2). There is evidence for significant gas motions, but in contrast to earlier results at low redshift, there are many sources in which the HI velocity is substantially negative (up to v=-1420km/s) with respect to the optical redshift, suggesting that in these sources the atomic gas, rather than falling into the centre, may be be flowing out, interacting with the jets, or rotating around the nucleus.Comment: 10 pages, accepted for publication in A&

A Supernova Factory in Mrk 273?

We report on 1.6 and 5.0 GHz observations of the ultraluminous infrared galaxy (ULIRG) Mrk 273, using the European VLBI Network (EVN) and the Multi-Element Radio-Linked Interferometer Network (MERLIN). We also make use of published 1.4 GHz VLBA observations of Mrk 273 by Carilli & Taylor (2000). Our 5 GHz images have a maximum resolution of 5-10 mas, which corresponds to linear resolutions of 3.5-7 pc at the distance of Mrk 273, and are the most sensitive high-resolution radio observations yet made of this ULIRG. Component N1, often pinpointed as a possible AGN, displays a steep spectral index ($\alpha = 1.2 \pm 0.1; S_\nu \propto \nu^{-\alpha}$); hence it is very difficult to reconcile with N1 being an AGN, and rather suggests that the compact nonthermal radio emission is produced by an extremely high luminous individual radio supernova (RSN), or a combination of unresolved emission from nested supernova remnants (SNR), luminous RSNe, or both. Component N2 is partly resolved out into several compact radio sources --none of which clearly dominates-- and a region of extended emission about 30 pc in size. The integrated spectral index of this region is flat ($\alpha = 0.15 \pm 0.1$), which can be interpreted as due to a superposition of several unresolved components, e.g., RSNe or SNRs, whose radio emission peaks at different frequencies and is partially free-free absorbed. The overall extended radio emission from component N is typical of nonthermal, optically thin radio emission ($\alpha = 0.8 \pm 0.1$), and its 1.4 GHz luminosity ($L_{1.4 \rm GHz} = (2.2 \pm 0.1)\times 10^{23}$ WHz$^{-1}$) is consistent with being produced by relativistic electrons diffused away from supernova remnants in an outburst.Comment: Accepted for publication in Monthly Notices of the Royal Astronomical Society Main Journal. 6 pages, 3 figure

3C459: A highly asymmetric radio galaxy with a starburst

Multifrequency radio observations of the radio galaxy 3C459 using MERLIN, VLA and the EVN, and an optical HST image using the F702W filter are presented. The galaxy has a very asymmetric radio structure, a high infrared luminosity and a young stellar population. The eastern component of the double-lobed structure is brighter, much closer to the nucleus and is significantly less polarized than the western one. This is consistent with the jet on the eastern side interacting with dense gas, which could be due to a merged companion or dense cloud of gas. The HST image of the galaxy presented here exhibits filamentary structures, and is compared with the MERLIN 5-GHz radio map. EVN observations of the prominent central component, which has a steep radio spectrum, show a strongly curved structure suggesing a bent or helical radio jet. The radio structure of 3C459 is compared with other highly asymmetric, Fanaroff-Riley II radio sources, which are also good candidates for studying jet-cloud interactions. Such sources are usually of small linear size and it is possible that the jets are interacting with clouds of infalling gas that fuel the radio source.Comment: 10 pages, 6 figures, 2 table

The puzzling case of the radio-loud QSO 3C 186: a gravitational wave recoiling black hole in a young radio source?

Context. Radio-loud AGNs with powerful relativistic jets are thought to be associated with rapidly spinning black holes (BHs). BH spin-up may result from a number of processes, including accretion of matter onto the BH itself, and catastrophic events such as BH-BH mergers. Aims. We study the intriguing properties of the powerful (L_bol ~ 10^47 erg s^-1) radio-loud quasar 3C 186. This object shows peculiar features both in the images and in the spectra. Methods. We utilize near-IR Hubble Space Telescope (HST) images to study the properties of the host galaxy, and HST UV and Sloan Digital Sky Survey optical spectra to study the kinematics of the source. Chandra X-ray data are also used to better constrain the physical interpretation. Results. HST imaging shows that the active nucleus is offset by 1.3 +- 0.1 arcsec (i.e. ~11 kpc) with respect to the center of the host galaxy. Spectroscopic data show that the broad emission lines are offset by -2140 +-390 km/s with respect to the narrow lines. Velocity shifts are often seen in QSO spectra, in particular in high-ionization broad emission lines. The host galaxy of the quasar displays a distorted morphology with possible tidal features that are typical of the late stages of a galaxy merger. Conclusions. A number of scenarios can be envisaged to account for the observed features. While the presence of a peculiar outflow cannot be completely ruled out, all of the observed features are consistent with those expected if the QSO is associated with a gravitational wave (GW) recoiling BH. Future detailed studies of this object will allow us to confirm this type of scenario and will enable a better understanding of both the physics of BH-BH mergers and the phenomena associated with the emission of GW from astrophysical sources.Comment: 16 pages, 8 figures. Accepted for publication in Astronomy & Astrophysics. New appendix adde

The Compact Structure of Radio-Loud Broad Absorption Line Quasars

We present the results of EVN+MERLIN VLBI polarization observations of 8 Broad Absorption Line (BAL) quasars at 1.6 GHz, including 4 LoBALs and 4 HiBALs with either steep or flat spectra on VLA scales. Only one steep-spectrum source, J1122+3124, shows two-sided structure on the scale of 2 kpc. The other four steep-spectrum sources and three flat-spectrum sources display either an unresolved image or a core-jet structure on scales of less than three hundred parsecs. In all cases the marginally resolved core is the dominant radio component. Linear polarization in the cores has been detected in the range of a few to 10 percent. Polarization, together with high brightness temperatures (from 2*10^9-5*10^10 K), suggest a synchrotron origin for the radio emission. There is no apparent difference in the radio orphologies or polarization between low-ionization and high-ionization BAL QSOs nor between flat- and steep-spectrum sources. We discuss the orientation of BAL QSOs with both flat and steep spectra, and consider a possible evolutionary scenario for BAL QSOs. In this scenario, BAL QSOs are probably the young population of radio sources, which are Compact Steep Spectrum or GHz peaked radio source analog at the low end of radio power.Comment: 16 pages, 3 figures, 4 tables; accepted for publication in MNRA