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PubMed1996

Simultaneous determination of isosorbide dinitrate and its mononitrate metabolites in human plasma by capillary gas chromatography with electron-capture detection.

A method for the simultaneous determination of isosorbide dinitrate (ISDN) and its mononitrate metabolites (2- and 5-ISMN) in human plasma by capillary gas chromatography with electron-capture detection was developed. Two internal standards were used: isomannide dinitrate (IMDN) for the determination of ISDN and isomannide mononitrate (IMMN) for the determinations of 2- and 5-ISMN. After addition of the internal standards, the compounds were isolated from plasma by solid-liquid extraction. They were determined by gas chromatography using an electron-capture detector. The reproducibility and accuracy of the method were found suitable in the range of concentrations 2.5-83 ng/ml for ISDN, 2.6-208 ng/ml for 2-ISMN and 2.3-1010 ng/ml for 5-ISMN. The limit of quantitation (LOQ) was about 2.5 ng/ml for each compound. The method was applied to clinical samples.

Chromatography, Gas↗

PubMed1996

Determination of 4-(4-chlorophenyl)-4-hydroxypiperidine, a metabolite of haloperidol, by gas chromatography with electron-capture detection.

An electron-capture gas chromatographic procedure was developed for the analysis of 4-(4-chlorophenyl)-4-hydroxypiperidine (CPHP), a metabolite of haloperidol. The assay involved basic extraction of this metabolite from the biological samples, followed by back-extraction with HCl. After basification of the acid phase, extractive derivatization with pentafluorobenzoyl chloride in toluene was conducted. The pentafluorobenzoyl derivative was quantified on a gas chromatograph equipped with a fused-silica capillary column, an electron-capture detector and a printer-integrator. N-(3-Trifluoromethylphenyl)piperazine was carried through the procedure as an internal standard and calibration curves were determined for each assay run. The procedure was demonstrated to be linear and reproducible and was utilized to detect and quantify CPHP in urine, plasma, brain and liver samples from rats treated with haloperidol. The structure of the derivatized metabolite was confirmed by gas chromatography-mass spectrometry.

Animals↗

PubMed1996

Analysis of 5-methoxytryptamine at the femtomole level in the rat and quail brain by gas chromatography-electron-capture negative-ion chemical ionization mass spectrometry.

A sensitive method for the measurement of endogenous 5-methoxytryptamine in brain tissue has been developed using capillary column gas chromatography-electron-capture negative-ion chemical ionization mass spectrometry. 5-Methoxytryptamine was first converted to N-[2H3]acetyl-5-methoxytryptamine by reaction with hexa-deuterated acetic anhydride, followed by reaction with pentafluoropropionic anhydride to yield the highly electron-capturing 3,3'-spirocyclic pentafluoro-propionyl indolenine derivative. Quantitative analysis was carried out by selected-ion monitoring of the [M-HF].- and [M-HF-DF].- ion intensity of the 3.3'-spirocyclic pentafluoropropionyl indolenine derivative, using 5-methoxy-[alpha, alpha, beta, beta-2H4]tryptamine as the internal standard. The presence of 5-methoxytryptamine in the brain tissue was demonstrated. In the absence of a monoamine oxidase inhibitor, the mean +/- S.D. levels of 5-methoxytryptamine in the rat and quail whole brain were found to be 30 +/- 6 and 347 +/- 52 pg/g, respectively. The possible physiological functions of 5-methoxytryptamine as a neuromodulator and/or neurotransmitter have to be considered.

5-Methoxytryptamine↗

PubMed1990

A review of techniques for the analysis of boron in the development of neutron capture therapy agents.

The resurgence of interest in neutron capture therapy, which uses boron-10 labelled compounds and neutron irradiation as a technique for the treatment of cancer, has been accompanied by the application of a new generation of techniques for the analysis and imaging of boron in biological matrices. In this review, a description is given of the requirements for boron neutron capture agents and the achievements to date of the analytical techniques.

Animals↗

PubMed1985

Plasma catecholamines and erythrocyte swelling following capture stress in a marine teleost fish.

Plasma concentrations of adrenaline and noradrenaline were measured at rest from cannulated fish and following net capture. Adrenaline and noradrenaline concentrations in capture-stressed fish averaged 36,740 pmol l-1 and 38,860 pmol l-1 respectively, whereas resting values were less than 200 pmol l-1 for both amines. Erythrocyte swelling and raised blood lactate were evident in stressed fish. In vitro effects of 5 mmol l-1 adrenaline on erythrocyte suspensions suggested that the catecholamine had a direct effect on erythrocyte volume. The significance of these results is discussed in relation to the oxygen transport properties of the blood.

Animals↗

PubMed1987

Boronation of antibodies with mercaptoundecahydro-closo-dodecaborate(2-) anion for potential use in boron neutron capture therapy.

The anionic polyhedral borane derivative, mercaptoundecahydro-closo-dodecaborate(2-), has been evaluated as a boronating agent for antibodies. The objective of these studies was the selective delivery of boron to neoplasms for neutron capture therapy. Incubation of a large excess of this anion with the polyclonal antibody antithymocyte globulin (ATG) resulted in the incorporation of 9-13 mol of the anion per mol of antibody. The extent of boron incorporation into the protein was measured either by tritium-labeled B12H11SH2- or by direct boron determination with neutron activation analysis. The nature of the covalent linkage of the anion to the antibody appeared to involve the formation of a new disulfide bond by a thiol-disulfide exchange. The number of boron atoms incorporated into antibodies by this method appeared to be inadequate for neutron capture therapy. However, such boronated antibodies may have potential for the detection of molecules of biologic interest by means of electron energy loss spectroscopy.

Antibodies, Monoclonal↗

PubMed1996

Loglinear models using capture-recapture methods to estimate the size of a measles epidemic.

Cases of measles reported separately by doctors, hospitals, laboratories, and others are treated as separate captures and releases of the population of children aged up to 10 years who have been infected by measles in a recent epidemic. Loglinear models are used to assess the adequacy of fit of various models for capture-recapture and the size of the epidemic in different age groups is estimated.

Australia↗

PubMedDate not supplied

Detection of IgM antibodies specific for measles virus by capture and indirect enzyme immunoassays.

During a measles outbreak, 112 serum specimens from 88 hospitalized patients were received in our laboratory for investigation of a morbilliform rash. These specimens (88 acute- and 24 convalescent-phase) were tested for the presence of measles-specific IgM antibodies by a capture EIA (enzyme immunoassay) using peroxidase-conjugated measles virus antigens and by an indirect EIA. Commercially available indirect EIA kits for measles-specific IgM antibodies were also used and compared with our homemade EIAs. Specificity studies included a collection of serum specimens containing rheumatoid factor, antinuclear antibodies or IgM antibodies specific to other viruses, and sera from blood donors and healthy children. Sensitivity of capture EIA and indirect EIA to detect measles IgM was 91.8 and 90.3%, respectively, and specificity was 98.2% for both tests. Specific IgM antibodies were detected in 70.5% of serum specimens at the first day after rash onset and were present for a month following the rash. Among the commercial measles IgM detection assays, EIA "Behring" was found to be a valid alternative for detection of measles virus-specific IgM.

Adolescent↗

PubMed1995

Development of an IgM capture assay for the diagnosis of B19 parvovirus infection using recombinant baculoviruses expressing VP1 or VP2 antigens.

BACKGROUND: The clinical manifestations of human parvovirus B19 infection are often similar to those induced as the result of infection by other infectious agents such as rubella and some bacteria. Although diagnosis of B19 infection is feasible by detection of specific antibodies, the tests require viraemic serum as a source of antigen. This inevitably leads to problems of reproducibility and dependence upon appropriate high quality clinical material. OBJECTIVES: To develop a monoclonal antibody capture ELISA (MACEIA) for detecting anti-B19 IgM antibody in human sera, using recombinant baculoviruses expressing the B19 parvovirus VP1 and VP2 proteins and to compare this with MACEIA using a plasma derived B19 antigen. STUDY DESIGN: Sera from 85 patients with proven B19 infection and the paired convalescent sera from 26 anti-B19 IgM-positive acute samples were examined for B19-specific IgM antibody by a monoclonal antibody capture assay that utilised recombinant baculoviruses expressing B19 proteins in lieu of a plasma-derived B19 antigen. Control samples consisted of 24 anti-rubella IgM, 24 anti-EBV IgM and 102 negative sera from uninfected individuals. RESULTS: Eighty-four of the 85 sera were anti-B19 IgM positive by MACEIA using recombinant baculovirus derived B19 antigen and by indirect immunofluorescence tests, whereas 79 were positive by MACEIA using plasma-derived antigen. Of the 26 convalescent samples which were positive as acute sera, 4 had become negative by 8 weeks post-infection. The expressed recombinant baculovirus antigens had identical molecular weights to the VP1 (84 kDa) and VP2 (58 kDa) proteins of virus purified from human plasma. Recombinant baculovirus-derived VP1 antigen was as effective as VP2 particles at detecting antibodies. CONCLUSIONS: Recombinant proteins VP1 and VP2, obtained from recombinant baculovirus-infected cell lysate, showed equal specificity to and higher sensitivity than, B19 virus purified from human plasma when used in MACEIA to detect B19-IgM antibody.

Journal Article↗

PubMed1994

A poliovirus type-specific IgM antibody-capture enzyme-linked immunosorbent assay for the rapid diagnosis of poliomyelitis.

OBJECTIVES: To develop and evaluate an IgM antibody-capture ELISA for the rapid detection of poliovirus type-specific IgM antibodies in serum and CSF of patients suspected of poliomyelitis. STUDY DESIGN: The assay uses monoclonal antibody to human IgM as catching antibody, monovalent IPV as antigen and type-specific monoclonal antibodies labeled with horseradish peroxidase as detecting antibody. RESULTS: Sera from 21 of 24 patients, contacts of these patients or recently vaccinated children were positive in the IgM assay. Sera from 100 healthy individuals were negative in all three poliovirus type-specific ELISAs. Sera from 5 of 81 patients with non-polio viral infections showed (weak) reactivity in one or more of the IgM assays. In a prospective study, sera and/or CSFs from 72 patients with a recent clinical diagnosis of poliomyelitis were tested. A homotypic IgM antibody response was found in 51 of the 55 patients which shedded wild poliovirus in stool or throat. Poliovirus type-specific IgM antibodies were also detected in serum from 13 of the 17 patients, from whom no poliovirus was isolated. Poliovirus type-specific IgM-antibodies could be detected from the second day up till at least 28 days after onset of paralysis. BACKGROUND: Poliovirus infections still create measurable morbidity and mortality in the world. Rapid diagnosis methods are needed to detect infection. CONCLUSION: The use of the poliovirus type-specific IgM antibody-capture ELISA allows rapid diagnosis of poliomyelitis within 24 h after obtaining serum or a CSF specimen.

Journal Article↗

PubMed1994

Estimating completeness of cancer registration in Saarland/Germany with capture-recapture methods.

Completeness of population-based cancer registration has been most commonly quantified by indirect measures, such as the death certificate only index or the mortality/incidence ratio. A major disadvantage of these measures is their strong dependence on the case fatality rate. Capture-recapture methodology offers an approach to estimate completeness directly which does not share this limitation. In this paper, a three-sources modelling approach is employed to derive estimates of completeness for the population-based cancer registry of Saarland. Overall, completeness is found to be high: estimates for all types of cancer range from 95.5 to 96.9% for calendar years 1970, 1975, 1980 and 1985. There is some variation with age (consistently high levels above age 30 years, a minimum of 87.7% in age group 15-29 years) and between cancer sites. Among the most common cancer sites, estimates of completeness are highest for gastrointestinal cancers (97.2%) and breast cancer (97.1%), while lower estimates of completeness are derived for cancers of the female genital organs (92.5%), the urinary tract (91.8%) and the prostate (91.0%). Although capture-recapture estimates are sensitive to the underlying assumptions about dependence between sources, careful application is encouraged to supplement traditional methods for evaluating completeness of cancer registration.

Adolescent↗

PubMed1992

Affinity capture electrophoresis for sequence-specific DNA purification.

A new method, affinity capture electrophoresis (ACE), has been developed for the sequence-specific isolation of DNA. The target DNA is complexed with a biotinylated probe and electrophoresed in a gel equipped with a trap of immobilized streptavidin. This selectively captures the target molecule and its biotinylated probe, while other nontarget molecules pass through the trap. The target DNA is subsequently recovered from the trap by destroying the interaction between the target DNA and the biotinylated probe. Two variations of this technique, one using triple-helix formation and the other using hybridization with a uracil-containing DNA probe at the end of the target fragment, proved effective in model experiments. Since this technique requires no denaturation and handles DNA inside an agarose gel matrix, it is, in principle, applicable to the isolation of very large DNAs.

Bacterial Proteins↗

PubMed2003

Detection and quantitation of fatty acid acyl conjugates of triamcinolone acetonide via gas chromatography-electron-capture negative-ion mass spectrometry.

The inherent electron-capture properties of triamcinolone acetonide (TAA) fatty acid conjugates were exploited for development of a GC-MS technique for quantitation of C21 long-chain fatty esters of TAA synthesized in BEAS-2B cells, an immortalized airway epithelium cell line. TAA esters extracted from BEAS-2B cells were purified and detected via selected ion monitoring of the molecular anions generated from the TAA esters under electron-capture negative-ion mass spectrometric conditions. Standard curves were linear over a range of 0.0 to >4.5 ng/mg protein with r(2) values = 1. Levels of TAA conjugates extracted from BEAS-2B treated with 10(-5)M TAA for 24h ranged from 0.024 to 0.301 ng/mg protein. Further evidence for confirmation of the identity of TAA fatty esters formed in BEAS-2B cells was obtained via selected reaction monitoring. The transition monitored was formation of the carboxy anion generated from each of the respective molecular anions of the TAA esters during collision-induced decomposition. These findings indicate that the GC-MS analysis is suitable for studies of the kinetics of the TAA fatty acid conjugates formation in vitro and may be directly applicable to determination of the kinetics of TAA fatty acid conjugation in vivo.

Anti-Infective Agents↗

PubMed2004

Cross-linking approach to affinity capture of protein complexes from chaotrope-solubilized cell lysates.

Affinity capture methods are widely used for isolation and analysis of protein complexes. Short peptide tags fused to the protein of interest normally facilitate straightforward purification and detection of interacting proteins. We investigated the suitability of applying C-terminally hexahistidine-tagged interleukin-12 (IL-12) alpha- and beta-chains as "bait" proteins for cocapturing novel binding partners using heterologous recombinant human embryonic kidney-293 (HEK-293) cell lines. The beta-chain, but not the alpha-chain, extracted from cell lysates was capable of binding to the Ni(2+)-nitrilotriacetic acid affinity resin under nondenaturing conditions. Retention of the alpha-chain on this matrix was dependent on treatment of cell lysates with high concentrations of chaotropes such as urea. Since under these conditions any noncovalent protein associations are destroyed, prior cross-linking of proteins interacting with the alpha-chain in intact cells was required. The use of the thiol-cleavable cross-linker 3,3'-dithiobis(succinimidyl proprionate) facilitated dissociation of alpha-chain-binding proteins by means of dithiothreitol following purification. Using this approach we were able to demonstrate a strong interaction between the endoplasmic reticulum chaperone calreticulin (CRT) and the IL-12 alpha-chain that was confirmed in a reciprocal anti-CRT immunoprecipitation assay. The assay presented here provides a simple approach to exposing concealed hexahistidine tags while retaining native noncovalent protein interactions and should be generally applicable in a range of pull-down or affinity capture methods aiming at analysis of protein complexes.

Affinity Labels↗

PubMed2005

Analysis of glycoprotein hormone receptor extracellular domain interactions using a solid-phase capture assay.

The receptors for the glycoprotein hormones are unique in having a large extracellular domain that is responsible for mediating ligand binding. We describe the characterization, validation, and application of a solid-phase radioligand binding assay that can be used to assess the interaction of peptides and small molecules at the extracellular domain (ECD) of the follicle-stimulating hormone receptor (FSHR). The assay utilizes a C-terminal tag on the FSHR-ECD, which is used to capture the ECD and position it in a sterically favorable orientation on a solid-phase platform. Competition experiments with the cognate ligand, FSH, indicated that the interaction at the FSHR-ECD using the solid-phase assay was comparable to the full-length receptor assayed using a standard filtration assay. The utility of the assay was evaluated by competing several peptides and a small molecule for both the full-length FSHR and the FSHR-ECD. The solid-phase capture format allowed for the establishment of an assay to specifically evaluate compounds that interact at the ECD or require the full-length receptor, thereby facilitating structure-activity studies. This assay format should be applicable to the other receptors of this family.

Binding, Competitive↗

PubMed2006

Detection and identification of Leishmania species in field-captured phlebotomine sandflies based on mini-exon gene PCR.

Leishmaniasis is one of the most diverse and complex of all vector-borne diseases. Because it involves several overlapping species and sandfly vectors, the disease has a complex ecology and epidemiology. Adequate therapy and follow-up depend on parasitological diagnosis, but classical methods present several constraints when identifying species. We describe a polymerase chain reaction (PCR) which uses primers designed from mini-exon repetitive sequences that are specific for subgenus LeishmaniaViannia (PV), as well as sequences with specificity for genus (PG) that can distinguish between Leishmania species from other insect flagellates with minor differences in PCR products. For standardization, these PCR were tested in experimentally infected sandflies, and Leishmania infection in these insects was successfully confirmed. This methodology identified a 3.9% infection rate in field-captured phlebotomine sandflies from an endemic region in Brazil. Natural infection by Leishmania species was identified in three samples of Lutzomyia longipalpis, of which two were Leishmania (L.) chagasi and one Leishmania (L.) amazonensis. Irrespective of specific epidemiological conclusions, the method used in this study was able to identify Leishmania infections both in experimentally infected and field-captured phlebotomine sandflies, and could be a useful tool in epidemiological studies and strategic planning for the control of human leishmaniasis.

Animals↗

PubMed2004

Effectiveness and safety of the Proxis system in demonstrating retrograde coronary blood flow during proximal occlusion and in capturing embolic material.

The Feasibility And Safety Trial for its embolic protection device during transluminal intervention in coronary vessels: a European Registry (FASTER) was designed to demonstrate that (1) the Proxis embolic protection system can control anterograde flow and reverse blood flow in native coronary arteries and saphenous vein grafts; and (2) this system can capture embolic debris. Percutaneous coronary intervention on stenotic coronary lesions revolutionized treatment of coronary disease, but is associated with the risk of major adverse cardiac events. This prospective, nonrandomized, multicenter clinical feasibility and safety study enrolled 40 patients with 51 lesions at 4 centers who underwent treatment of stenotic lesions with proximal emboli protection (Proxis system). Proxis was successfully used 95% of the time, and embolic debris was qualitatively identified in all cases. Major adverse cardiac events occurred in 2 patients (5.0%): 1 late in-stent thrombosis resulting in death and 1 non-Q-wave infarction when a lesion was crossed before deployment of the Proxis system. Mean vessel occlusion time was 4.3 +/- 2.4 minutes. Native flow reversal was sufficient in 31 patients, with a mean aspirate volume of 11.8 +/- 6.5 ml. When the infusion catheter was used to augment reflow, mean occlusion time was 4.6 minutes. In conclusion, this trial is the first to demonstrate that retrograde blood flow can be achieved during proximal occlusion and that the Proxis system can be used safely during intervention of saphenous vein grafts and native coronary arterial lesions to capture embolic material.

Aged↗

PubMed2006

Absolute activity measurement of the electron-capture-based radionuclides 139Ce, 125I, 192Ir and 65Zn by liquid scintillation coincidence counting.

Four radionuclides with electron-capture-based decay schemes have been directly measured by a liquid scintillation coincidence extrapolation technique. 125I, 192Ir and 65Zn were measured as part of international key comparisons held under the auspices of the International Bureau of Weights and Measures (BIPM). The 139Ce measurements formed part of a regional comparison organized by the Asia Pacific Metrology Programme (APMP). Since 139Ce decays purely by electron-capture, the basic method is described for this radionuclide. Results and difficulties encountered are discussed and uncertainty budgets are presented.

Algorithms↗