A cryogenic ice setup to simulate carbon atom reactions in interstellar ices.

The design, implementation, and performance of a customized carbon atom beam source for the purpose of investigating solid-state reaction routes in interstellar ices in molecular clouds are discussed. The source is integrated into an existing ultrahigh vacuum setup, SURFace REaction SImulation DEvice (SURFRESIDE2), which extends this double atom (H/D, O, and N) beamline apparatus with a third atom (C) beamline to a unique system that is fully suited to explore complex organic molecule solid-state formation under representative interstellar cloud conditions. The parameter space for this system is discussed, which includes the flux of the carbon atoms hitting the ice sample, their temperature, and the potential impact of temperature on ice reactions. Much effort has been put into constraining the beam size to within the limits of the sample size with the aim of reducing carbon pollution inside the setup. How the C-atom beam performs is quantitatively studied through the example experiment, C + 18O2, and supported by computationally derived activation barriers. The potential for this source to study the solid-state formation of interstellar complex organic molecules through C-atom reactions is discussed.

Spectroscopic constraints on CH3OH formation: CO mixed with CH3OH ices towards young stellar objects.

The prominent infrared absorption band of solid CO - commonly observed towards young stellar objects (YSOs) - consists of three empirically determined components. The broad 'red component' (2136 cm-1, 4.681 µm) is generally attributed to solid CO mixed in a hydrogen-bonded environment. Usually, CO embedded in the abundantly present water is considered. However, CO:H2O mixtures cannot reproduce the width and position of the observed red component without producing a shoulder at 2152 cm-1, which is not observed in astronomical spectra. Cuppen et al. showed that CO:CH3OH mixtures do not suffer from this problem. Here, this proposition is expanded by comparing literature laboratory spectra of different CO-containing ice mixtures to high-resolution (R = λ/Δλ = 25000) spectra of the massive YSO AFGL 7009S and of the low-mass YSOL1489 IRS. The previously unpublished spectrum of AFGL 7009S shows a wide band of solid 13CO, the first detection of 13CO ice in the polar phase. In this source, both the 12CO and 13CO ice bands are well fitted with CO:CH3OH mixtures, while respecting the profiles and depths of the methanol bands at other wavelengths, whereas mixtures with H2O cannot. The presence of a gradient in the CO:CH3OH mixing ratio in the grain mantles is also suggested. Towards L1489 IRS, the profile of the 12CO band is also better fitted with CH3OH-containing ices, although the CH3OH abundance needed is a factor of 2.4 above previous measurements. Overall, however, the results are reasonably consistent with models and experiments about formation of CH3OH by the hydrogenation of CO ices.

THz and mid-IR spectroscopy of interstellar ice analogs: methyl and carboxylic acid groups.

A fundamental problem in astrochemistry concerns the synthesis and survival of complex organic molecules (COMs) throughout the process of star and planet formation. While it is generally accepted that most complex molecules and prebiotic species form in the solid phase on icy grain particles, a complete understanding of the formation pathways is still largely lacking. To take full advantage of the enormous number of available THz observations (e.g., Herschel Space Observatory, SOFIA, and ALMA), laboratory analogs must be studied systematically. Here, we present the THz (0.3-7.5 THz; 10-250 cm(-1)) and mid-IR (400-4000 cm(-1)) spectra of astrophysically-relevant species that share the same functional groups, including formic acid (HCOOH) and acetic acid (CH3COOH), and acetaldehyde (CH3CHO) and acetone ((CH3)2CO), compared to more abundant interstellar molecules such as water (H2O), methanol (CH3OH), and carbon monoxide (CO). A suite of pure and mixed binary ices are discussed. The effects on the spectra due to the composition and the structure of the ice at different temperatures are shown. Our results demonstrate that THz spectra are sensitive to reversible and irreversible transformations within the ice caused by thermal processing, suggesting that THz spectra can be used to study the composition, structure, and thermal history of interstellar ices. Moreover, the THz spectrum of an individual species depends on the functional group(s) within that molecule. Thus, future THz studies of different functional groups will help in characterizing the chemistry and physics of the interstellar medium (ISM).

Solid state chemistry of nitrogen oxides--part I: surface consumption of NO.

The role of nitrogen and oxygen chemistry in the interstellar medium is still rather poorly understood. Nitric oxide, NO, has been proposed as an important precursor in the formation of larger N- and O-bearing species, such as hydroxylamine, NH2OH, and nitrogen oxides, NO2 and N2O. The topic of this study is the solid state consumption of NO via oxygenation and the formation of NO2 and other nitrogen oxides (ONNO2 and N2O4) under conditions close to those encountered on icy grains in quiescent interstellar clouds. In our experiments nitric oxide and oxygen allotropes (O, O2, and O3) or N atoms are co-deposited under ultra-high vacuum conditions on different substrates (silicate, graphite, compact ASW ice, and gold) at temperatures ranging between 10 and 35 K. Reaction products are monitored via Fourier Transform Reflection Absorption Infrared Spectroscopy (FT-RAIRS) and Temperature Programmed Desorption (TPD) using mass spectrometry. We find that NO2 is efficiently formed in NO + O/O2/O3/N solid surface reactions. These are essentially barrier free and offer a pathway for the formation of NO2 in space. Nitrogen dioxide, however, has not been astronomically detected, contradicting the efficient reaction channel found here. This is likely due to other pathways, including regular hydrogenation reactions, as discussed separately in part II of this study.

Solid state chemistry of nitrogen oxides--part II: surface consumption of NO2.

Nitrogen oxides are considered to be important astrochemical precursors of complex species and prebiotics. However, apart from the hydrogenation of solid NO that leads to the surface formation of hydroxylamine, little is known about the full solid state reaction network involving both nitrogen and oxygen. Our study is divided into two papers, hereby called Part I and Part II. In the accompanying paper, we investigate the surface reactions NO + O/O2/O3 and NO + N with a focus on the formation of NO2 ice. Here, we complement this study by measurements of the surface destruction of solid NO2, e.g., NO2 + H/O/N. Experiments are performed in two separate ultra-high vacuum setups and therefore under different experimental conditions to better constrain the experimental results. Surface reaction products are monitored by means of Fourier Transform Reflection Absorption Infrared Spectroscopy (FT-RAIRS) and Temperature Programmed Desorption (TPD) techniques using mass spectrometry. The surface destruction of solid NO2 leads to the formation of a series of nitrogen oxides such as NO, N2O, N2O3, and N2O4 as well as HNO, NH2OH, and H2O. When NO2 is mixed with an interstellar more relevant apolar (i.e., CO) ice, solid CO2 and HCOOH are also formed due to interactions between different reaction routes. The astrophysical implications of the full nitrogen and oxygen reaction network derived from Parts I and II are discussed.

SURFRESIDE(2): an ultrahigh vacuum system for the investigation of surface reaction routes of interstellar interest.

A new ultrahigh vacuum experiment is described to study atom and radical addition reactions in interstellar ice analogues for astronomically relevant temperatures. The new setup - SURFace REaction SImulation DEvice (SURFRESIDE(2)) - allows a systematic investigation of solid state pathways resulting in the formation of molecules of astrophysical interest. The implementation of a double beam line makes it possible to expose deposited ice molecules to different atoms and/or radicals sequentially or at the same time. Special efforts are made to perform experiments under fully controlled laboratory conditions, including precise atom flux determinations, in order to characterize reaction channels quantitatively. In this way, we can compare and combine different surface reaction channels with the aim to unravel the solid state processes at play in space. Results are constrained in situ by means of a Fourier transform infrared spectrometer and a quadrupole mass spectrometer using reflection absorption infrared spectroscopy and temperature programmed desorption, respectively. The performance of the new setup is demonstrated on the example of carbon dioxide formation by comparing the efficiency through two different solid state channels (CO + OH → CO2 + H and CO + O → CO2) for which different addition products are needed. The potential of SURFRESIDE(2) to study complex molecule formation, including nitrogen containing (prebiotic) compounds, is discussed.

Efficient surface formation route of interstellar hydroxylamine through NO hydrogenation. II. The multilayer regime in interstellar relevant ices.

Hydroxylamine (NH(2)OH) is one of the potential precursors of complex pre-biotic species in space. Here, we present a detailed experimental study of hydroxylamine formation through nitric oxide (NO) surface hydrogenation for astronomically relevant conditions. The aim of this work is to investigate hydroxylamine formation efficiencies in polar (water-rich) and non-polar (carbon monoxide-rich) interstellar ice analogues. A complex reaction network involving both final (N(2)O, NH(2)OH) and intermediate (HNO, NH(2)O·, etc.) products is discussed. The main conclusion is that hydroxyl-amine formation takes place via a fast and barrierless mechanism and it is found to be even more abundantly formed in a water-rich environment at lower temperatures. In parallel, we experimentally verify the non-formation of hydroxylamine upon UV photolysis of NO ice at cryogenic temperatures as well as the non-detection of NC- and NCO-bond bearing species after UV processing of NO in carbon monoxide-rich ices. Our results are implemented into an astrochemical reaction model, which shows that NH(2)OH is abundant in the solid phase under dark molecular cloud conditions. Once NH(2)OH desorbs from the ice grains, it becomes available to form more complex species (e.g., glycine and ß-alanine) in gas phase reaction schemes.

Water formation by surface O3 hydrogenation.

Three solid state formation routes have been proposed in the past to explain the observed abundance of water in space: the hydrogenation reaction channels of atomic oxygen (O + H), molecular oxygen (O(2) + H), and ozone (O(3) + H). New data are presented here for the third scheme with a focus on the reactions O(3) + H, OH + H and OH + H(2), which were difficult to quantify in previous studies. A comprehensive set of H/D-atom addition experiments is presented for astronomically relevant temperatures. Starting from the hydrogenation/deuteration of solid O(3) ice, we find experimental evidence for H(2)O/D(2)O (and H(2)O(2)/D(2)O(2)) ice formation using reflection absorption infrared spectroscopy. The temperature and H/D-atom flux dependence are studied and this provides information on the mobility of ozone within the ice and possible isotope effects in the reaction scheme. The experiments show that the O(3) + H channel takes place through stages that interact with the O and O(2) hydrogenation reaction schemes. It is also found that the reaction OH + H(2) (OH + H), as an intermediate step, plays a prominent (less efficient) role. The main conclusion is that solid O(3) hydrogenation offers a potential reaction channel for the formation of water in space. Moreover, the nondetection of solid ozone in dense molecular clouds is consistent with the astrophysical picture in which O(3) + H is an efficient process under interstellar conditions.

Water formation at low temperatures by surface O2 hydrogenation I: Characterization of ice penetration.

Water is the main component of interstellar ice mantles, is abundant in the solar system and is a crucial ingredient for life. The formation of this molecule in the interstellar medium cannot be explained by gas-phase chemistry only and its surface hydrogenation formation routes at low temperatures (O, O(2), O(3) channels) are still unclear and most likely incomplete. In a previous paper we discussed an unexpected zeroth-order H(2)O production behavior in O(2) ice hydrogenation experiments compared to the first-order H(2)CO and CH(3)OH production behavior found in former studies on hydrogenation of CO ice. In this paper we experimentally investigate in detail how the structure of O(2) ice leads to this rare behavior in reaction order and production yield. In our experiments H atoms are added to a thick O(2) ice under fully controlled conditions, while the changes are followed by means of reflection absorption infrared spectroscopy (RAIRS). The H-atom penetration mechanism is systematically studied by varying the temperature, thickness and structure of the O(2) ice. We conclude that the competition between reaction and diffusion of the H atoms into the O(2) ice explains the unexpected H(2)O and H(2)O(2) formation behavior. In addition, we show that the proposed O(2) hydrogenation scheme is incomplete, suggesting that additional surface reactions should be considered. Indeed, the detection of newly formed O(3) in the ice upon H-atom exposure proves that the O(2) channel is not an isolated route. Furthermore, the addition of H(2) molecules is found not to have a measurable effect on the O(2) reaction channel.

Water formation at low temperatures by surface O2 hydrogenation II: The reaction network.

Water is abundantly present in the Universe. It is the main component of interstellar ice mantles and a key ingredient for life. Water in space is mainly formed through surface reactions. Three formation routes have been proposed in the past: hydrogenation of surface O, O(2), and O(3). In a previous paper [Ioppolo et al., Astrophys. J., 2008, 686, 1474] we discussed an unexpected non-standard zeroth-order H(2)O(2) production behaviour in O(2) hydrogenation experiments, which suggests that the proposed reaction network is not complete, and that the reaction channels are probably more interconnected than previously thought. In this paper we aim to derive the full reaction scheme for O(2) surface hydrogenation and to constrain the rates of the individual reactions. This is achieved through simultaneous H-atom and O(2) deposition under ultra-high vacuum conditions for astronomically relevant temperatures. Different H/O(2) ratios are used to trace different stages in the hydrogenation network. The chemical changes in the forming ice are followed by means of reflection absorption infrared spectroscopy (RAIRS). New reaction paths are revealed as compared to previous experiments. Several reaction steps prove to be much more efficient (H + O(2)) or less efficient (H + OH and H(2) + OH) than originally thought. These are the main conclusions of this work and the extended network concluded here will have profound implications for models that describe the formation of water in space.

Long-term serological evaluation of patients with cystic echinococcosis treated with benzimidazole carbamates.

Seeking better immunological markers indicating the long-term outcome of cystic echinococcosis (CE) after chemotherapy we studied 23 patients receiving albendazole, clinically followed for 8 years, and grouped ultrasonographically according to therapeutic outcome. Antibody responses against a partially purified fraction of hydatid fluid (HFF) and antigen B (AgB) were evaluated by indirect haemagglutination (IHA), ELISA and immunoblotting (IB). Although IHA titres varied over the course of treatment, differences in mean antibody titres to HFF between groups were significant only at 4 years (P = 0.031). IgG isotype expression remained unchanged during follow-up whereas IgE expression decreased in patients with cured or stable disease. AgB disclosed higher IgG4 expression (P < 10(-4); P = 0.025) and lower IgG1 expression than HFF (P < 10(-4); P = 0.022). IHA antibody titres were higher in patients with progressive than in those with cured or stable disease, even in those with the same cyst type. ELISA isotype profiles differed between groups, particularly for type CE 3, 4 and 5 cysts: higher serum IgG1 and IgG3, lower IgG4 and IgE in patients with cured or stable disease. Although combined serological testing provides scarce information on the long-term outcome of CE after chemotherapy it may be useful for reviewing in a retrospective study the outcome of a cyst and for assessing the host-parasite relationship.

Modulation of human immune response by Echinococcus granulosus antigen B and its possible role in evading host defenses.

By directly suppressing the function of certain immune cell subsets and by stimulating other cell populations related to immunopathology, parasite-derived substances play an important role in the chronic establishment of parasitic disease. Our objective was twofold: (i) to investigate further the role of Echinococcus granulosus antigen B (AgB) in the human early inflammatory response by determining its effect on polymorphonuclear cell (PMN) random migration, chemotaxis, and oxidative metabolism and (ii) to determine its action in acquired immunity by evaluating AgB and sheep hydatid fluid (SHF)-driven Th1 (gamma interferon [IFN-gamma] and interleukin 12 [IL-12]) and Th2 (IL-4 and IL-13) cytokine production by peripheral blood mononuclear cells (PBMC) from 40 patients who had cured or stable or progressive cystic echinococcosis. AgB significantly inhibited PMN recruitment but left their random migration and oxidative metabolism unchanged. Patients' PBMC stimulated with AgB produced IL-4 and IL-13 but did not produce IL-12. They also produced significantly lower IFN-gamma concentrations than did PBMC stimulated with SHF (P = 10(-5)). AgB skewed the Th1/Th2 cytokine ratios towards a preferentially immunopathology-associated Th2 polarization, predominantly in patients with progressive disease. AgB-stimulated patients' PBMC also proliferated less than SHF-stimulated PBMC (P = 9 x 10(-3)). In vitro Th2 cytokine production was reflected in vivo by elevated specific immunoglobulin E (IgE) and IgG4 antibodies binding to AgB. These findings confirm that AgB plays a role in the escape from early immunity by inhibiting PMN chemotaxis. They also add new information on the host-parasite relationship, suggesting that AgB exploits the activation of T helper cells by eliciting a nonprotective Th2 cell response.

Native and recombinant antigens in the immunodiagnosis of human cystic echinococcosis.

To evaluate the diagnostic sensitivity and specificity of immunoelectrophoresis (IEP), indirect haemagglutination (IHA), enzyme-linked immunosorbent assay (ELISA) and immunoblotting (IB), we compared their ability in detecting IgG antibodies to a hydatid fluid fraction (HFF) and to native and recombinant antigen B of Echinococcus granulosus. We tested sera from patients who had cystic echinococcosis (CE) grouped according to their type of cysts (n = 204), from patients with other parasitic diseases (n = 21), lung or liver carcinomas (n = 6) or serous cysts (n = 26) and from healthy controls (n = 90). HFF-IB gave the highest sensitivity (80%) followed by ELISA (72%), IHA (54%) and IEP (31%), respectively. The diagnostic sensitivity significantly (P < 0.01) decreased as cysts matured from type I-II to type VII. Recombinant and native antigen B-IB yielded similar sensitivity (74%). A large number of clinically or surgically confirmed CE patients (20%) resulted negative. In these patients' sera, IB to assess the usefulness of the recombinant E. granulosus elongation factor-1 beta/delta in detecting IgE antibodies yielded 33% of positivity. Our findings underline the need to standardize techniques and antigenic preparations and to improve the performance of immunodiagnosis by characterizing new antigens and detecting distinct immunoglobulin classes.

Immunochemical and molecular characterization of vertebral hydatid fluid.

This study describes a 42-year-old male born in Tunisia, who presented with vertebral hydatidosis which was initially misdiagnosed as Pott's disease. The correct diagnosis was finally made utilizing CT and MRI imaging and confirmation of the diagnosis was made possible by molecular and immunochemical characterization of the content of vertebral cyst fluid.

Serum cytokine detection in the clinical follow up of patients with cystic echinococcosis.

The relation of serum cytokine levels and outcome of chemotherapy was evaluated in 15 patients with cystic echinococcosis. Serum IL-4, IL-10 and interferon-gamma (IFN-gamma) concentrations were determined by ELISA before and after a 3-month course of albendazole treatment: at least one serum sample per patient from 13 patients (87%) contained measurable amounts of IL-4; samples from five patients (33%) measurable amounts of IL-10 and samples from only two patients (13%) measurable amounts of IFN-gamma. Clinical assessment at 1 year after the end of therapy showed that 11 of the 15 patients had responded clinically. Seven of these patients had lower IL-4 serum concentrations, two had unchanged and two undetectable amounts (pre- versus post-therapy, n = 11, P = 0.008). Conversely, of the patients who did not respond, three had higher and one patient unchanged serum IL-4 concentrations. Serum IL-10 10 levels also decreased in all patients who responded (3/5) and increased in all patients who did not (2/5). No association was found between cytokine concentrations and cyst characteristics or antibody levels. Overall these data suggest that serum IL-4 detection may be useful in the follow up of patients with cystic echinococcosis.

Cytokine patterns in seropositive and seronegative patients with Echinococcus granulosus infection.

To investigate the involvement of immunological variables, including the Th1/Th2 balance, in the mechanisms underlying seronegativity in cystic echinococcosis (CE), Th1 and Th2 cytokines were evaluated in ELISA determining IFN-gamma, IL-4, -5 and -10 production in peripheral blood mononuclear cell (PBMC) cultures from 16 patients (six seronegative and ten seropositive) with CE and from six healthy donors. Patients who had undetectable antibody levels in their sera invariably produced no IL-5 and scarce IL-4 and -10 (Th2 response); both groups of patients produced similar amounts of IFN-gamma; (Th1 response). The analysis of host clinical features and cyst characteristics shows that these variables do not explain the lack of detectable serum antibody levels. These findings suggest that seronegativity arises because unknown host or parasite factors, or both, cause inadequate Th2 cell activation thus reducing production of Th2 cytokines, including IL-5, the cytokine primarily implicated in immunoglobulin expression in CE.

Immunological responses to antigen B from Echinococcus granulosus cyst fluid in hydatid patients.

The immunological reactivity of Echinococcus granulosus antigen B was evaluated in 30 hydatid patients. Antigen B was purified from sheep hydatid cyst fluid by electroelution from a non-reducing SDS-PAGE gel (AgB). In ELISA and immunoblotting (IB), determining antibody production in sera from patients with hydatid disease and with other parasitic infections, purified AgB showed higher specificity than a partially purified antigen named pH5PPT (100% vs 83% in pH5PPT-ELISA and 58% in pH5PPT-IB). AgB-IB achieved higher sensitivity than AgB-ELISA (80% vs 63%). All AgB-IB positive sera recognized the 12 kDa subunit. Qualitative AgB-IB assessment of IgG isotype responses identified IgG4 as the predominant isotype (87%). The other isotypes showed a lower percentage of positive reactions: IgG1, 33%; IgG2, 21%; and IgG3, 17%. PBMC proliferative assay revealed a cellular response to AgB in 100% of patients' PBMC. These findings confirm antigen B, especially its smallest subunit, as a good diagnostic molecule.

Immunological markers indicating the effectiveness of pharmacological treatment in human hydatid disease.

The relation of interferon-gamma (IFN-gamma), IL-4, IL-10 production and specific IgE, total IgG, IgG subclass expression to the effectiveness of pharmacological treatment in human hydatid disease (Echinococcus granulosus infection) was evaluated in 27 hydatid patients divided into four clinical groups according to their response to albendazole/mebendazole therapy (full, partial, low and non-responders). After parasite antigen stimulation, peripheral blood mononuclear cells (PBMC) from full responders produced significantly more IFN-gamma (P = 0.038), significantly less IL-4 (P = 0.001) and less IL-10 than PBMC from non-responders. PBMC from partial and low responders produced intermediate cytokine concentrations. ELISA determining immunoglobulin production showed that sera from all non-responders had IgE and IgG4 antibodies, both regulated by IL-4. In contrast to IgG4, IgE decreased rapidly in full responders. Full responders also showed the highest percentage of IgG3 reactions. Qualitative analysis of total IgG responses in hydatid patients' sera determined by immunoblotting showed that binding profiles to hydatid cyst fluid antigen differed in the four groups of treated patients. Non-responders had the highest percentage of reactions to all subunits of antigens 5 and B, and full responders had the highest percentage of reactions to antigen 5 alone. The high IFN-gamma production associated with a lack of IL-4 and low IL-10 production in the full responders, and vice versa the high IL-4 and IL-10 production associated with lack of or low IFN-gamma production in the non-responders implies Th1 cell activation in protective immunity and Th2 cell activation in susceptibility to hydatid disease. IgE may be a useful marker of therapeutic success in hydatid patients with pretreatment specific IgE antibodies. IgG subclass responses and differential immunoglobulin subclass binding pattern to hydatid antigens may also be useful in the immunosurveillance of hydatid disease.

Cross-linking of at least three binding sites mediates signal transduction in a CR2-positive Burkitt lymphoma derived cell line (Raji).

In this study we demonstrate that Raji cells, a CR2-positive Burkitt lymphoma-derived cell line, during cell growth, need the cross-linking of multiple OKB7 binding sites or C3d determinants to mediate signal transduction. The loss of one of these affects the cellular response. Moreover, OKB7, the anti-CR2 MoAb, recognizes C3d determinants on the cell surface and inhibits signal transduction induced by anti-C3d polyclonal antibody. Since Raji cells are always CR2 positive during cell growth, we suppose that at least another protein, along with CR2, may be involved in setting up a cell surface complex able to receive and transduce the signal triggered by OKB7. In our experimental system the protein that offers a third binding site to OKB7, may be represented by a 33 kDa protein bearing C3d determinants.

Use of a monoclonal antibody against the antigen B of Echinococcus granulosus for purification and detection of antigen B.

A monoclonal antibody directed against the 8 kDa subunit of antigen B of Echinococcus granulosus was raised. This antibody was used for purification of antigen B by affinity chromatography. SDS-PAGE and immunological analysis of the purified antigen B demonstrated that the 8 kDa antigen B subunit was purified to homogeneity. The purified antigen retained its strong immunoreactivity in ELISA using human hydatid sera. Furthermore, a sandwich ELISA was established for detection of antigen B from hydatid cysts. The usefulness of this test system was demonstrated by the detection of antigen B in human hydatid cyst fluids, thus confirming the diagnosis of an echinococcal disease.