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AGENE in bread.

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Bread↗

[Detection of bovine herpesvirus-1 using the dot-blot hybridization method].

With the development of molecular biology in the 1980s methods of microorganism detection start to innovate. One of the main advantages of the molecular-genetic methods, namely hybridization of nucleic acids and PCR methods, is the detection of genome of microorganism without the need for cellular cultivation. To detect BHV-1 (etiological agens IBR-IPV) the dot-blot hybridization method on nitrocellulose filters was used together with different types of DNA probes (two-fiber recombinant plasmids, one-fiber recombinant phages M 13 and 40 bp synthetic oligonucleotide). Genome DNA BHV-1 was isolated from samples (virions, infested cells, nasal smears and secretions by phenol extraction). The highest sensitivity of detection was achieved with 32P-pUR-1 probe (1.8kb random EcoRI-Hind III fragment ligated into plasmid pUC 9) which detected genome BHV-1 in 5 x 10(3) infested MDBK cells. This probe did not respond with herpetic viruses BHV-2, BHV-3, BHV-4 and the virus of Aujeszky's disease. The quality of pUR-1 probe was further tested for IBR diagnostics in animals experimentally infested with the virus BHV-1 (intranasal infection). BHV-1 could be detected in nasal smears and secretions in experimentally infested calves as early as on the first day following infection, while the agens amount reached its peak on the days 2-3 and on the days 6-7 the occurrence of virus fell markedly. When digoxigenin-pUR-1, i.e. non-radioactively marked probe, the virus presence was confirmed only on the days 2-3, in the time of the highest occurrence of infection agens. To detect the virus through the dot-blot hybridization nasal secretions were confirmed as better compared with nasal smears. The technology of virus isolation on cell cultures confirmed also the occurrence of agens as soon as on the first day from infection, with maximum on the days 2-5, but much more reliably it detected the virus on the days observed from the day 3 and their peak was obtained on the day 6 from infection. Experiments, comparing classical methods of IBR diagnostics (detection of specific antibodies, the method of isolation on cellular cultures) with the dot-blot hybridization using the samples obtained from farms with natural occurrence of IBM, are under progress.

Animals↗

An evaluation of an enzyme immunoassay method for immunoreactive trypsin in dried blood spots.

A novel monoclonal antibody based enzyme immunoassay (EIA) method for the measurement of the human cationic trypsinogen (NeoScreen, AGEN Biomedical Ltd., Acacia Ridge, Australia) in dried blood spots for the neonatal screening of cystic fibrosis was evaluated. The calibration standards provided as dried blood spots by AGEN are highly unstable and must be replaced with user prepared materials. Reference values from control individuals were obtained by parametric methods. A preliminary comparison with a polyclonal antibody based RIA method (Trypsik, SORIN Biomedica, Saluggia, Italy) was performed. Regression analysis between the RIA and the EIA methods gave a coefficient of correlation of 0.58 for RIA values less than 40 micrograms/L and of 0.77 for RIA values greater than or equal to 40 micrograms/L. Average CV of the within-run imprecision for the EIA method was 19.6% and for the RIA method 28.8%. CVs of the between-run imprecision at low, intermediate and high values for the EIA method were 23.7%, 15.8%, 15.6% and for the RIA method 20.6%, 14.4%, 11.2%. The diagnostic accuracy analyzed by a Receiver Operating Characteristics (ROC) curve of the RIA method gave a maximum accuracy of 190.9 while that of a simulated ROC curve for the EIA method was 193.0. We found that the precision and the diagnostic accuracy of the EIA method (AGEN) are equal to or better than those of one of the RIA methods.

Calibration↗

Agenesis of the corpus callosum: limits of functional compensation.

Two patients with radiologically confirmed total agenesis of the corpus callosum, and free of gross focal hemispheric pathology, received a battery of lateralized and free-field language and perceptual-motor tests. These tasks allowed a comparison of (1) agenic findings with previous results from surgical commissurotomy patients, and (2) the inter- and intrahemispheric performance for each agenic subject. In general, the results indicate that the two agenic patients do not demonstrate the marked deficits of cross-integration reported after commissurotomy. However, there was an upper limit to their integrative capacity for visual, and to an even greater extent, for fine motor and kinesthetically mediated performance. The tasks that proved particularly difficult appeared to have common factors which may be related to functional capabilities of the ipsilateral motor pathways and anterior commissure.

Adult↗

Methionine sulfoximine shows excitotoxic actions in rat cortical slices.

Methionine sulfoximine (MSO) is a rare amino acid. It occurs in nature or as a by-product of some forms of food processing. A notable example of the latter was a former method for bleaching wheat flour, using nitrogen trichloride, the "agene process," in use for most of the first 50 years of this century. "Agenized" flour was found to be responsible for various neurological disorders in animals, and MSO was identified as the toxic factor. The agene process was subsequently discontinued in the United States and the United Kingdom circa 1950. MSO inhibits the synthesis of both glutathione and glutamine, and it is possible that its actions on the nervous system arise from alterations in the amount or distribution of these molecules. Structurally, MSO resembles glutamate, an observation that has also raised the possibility that it might have more direct glutamate-like actions on neurons. In the present investigation, we report excitatory and toxic actions of MSO in an in vitro preparation of adult rat cortex. Field potential recordings in this preparation show that MSO application evokes a sustained depolarization, which can be blocked by the N-methyl-D-aspartate (NMDA) antagonist L-(+)-2-amino-5-phosphonovalerate (AP5). However, competition assays using MSO on [3H]CGP-39653 (DL-(E)-2-amino-4-propyl-1-phosphono-3-pentenoate) binding in rat cortical homogenates show only 20% displacement of total binding, suggesting that MSO is acting indirectly, perhaps by releasing glutamate. To investigate this possibility, we measured glutamate release during MSO application. Time course and dose-response experiments with MSO showed significant [3H]glutamate release, which was partially attenuated by AP5. To assess cellular toxicity, we measured lactate dehydrogenase (LDH) release from cortical sections exposed to MSO. MSO treatment led to a rapid increase in LDH activity, which could be blocked by AP5. These data suggest that MSO acts by increasing glutamate release, which then activates NMDA receptors, leading to excitotoxic cell death. These data suggest the possibility that MSO in processed flour had excitotoxic actions that may have been contributing factors to some human neuronal disorders.

2-Amino-5-phosphonovalerate↗

[Uptake and distribution of Camposan in rye].

Experiments for uptaking and distribution of the culm stabiliser "camposan" with the agens ethephon are very important to tell something about the dwarf behaviour of the treated plants of rye. The radioactive labelled ethephon is infiltrated through the roots, leaves and cuttings of culms. Uptaking through the roots is more complete than those about the foliar dissepiment. Otherwise uptaking through the foliar dissepiment is higher than about the leaves only. After radioautographic experiments, culmcutting-experiments and experiments with intact rye in the 5--6 leaf-stage we have found that the agens is transported acropetal in the xylem of the plants after uptaking by the roots up to 3 days. The labelled ethephon is translocated in a higher concentration in the phloem after foliar dissepiment application. The concentration of the agens decreases in the investigated plants up to the 5th day enormously (40--50% and after then slightly following the degradation to gaseous compounds. During 4 days ethephon is metabolized to about 5--15% ethylene and additional to about 18--20% carbondioxide.

Autoradiography↗

Longitudinal analysis of deciduous tooth emergence: II. Parametric survival analysis in Bangladeshi, Guatemalan, Japanese, and Javanese children.

We present a form of parametric survival analysis that incorporates exact, interval-censored, and right-censored times to deciduous tooth emergence. The method is an extension of common cross-sectional procedures such as logit and probit analysis, so that data arising from mixed longitudinal and cross-sectional studies can be properly combined. We extended the method to incorporate and estimate a proportion of agenic teeth. While we concentrate on deciduous tooth emergence, the method is relevant to studies of permanent tooth emergence and other developmental events. Deciduous tooth emergence data were analyzed from four longitudinal studies. The samples are 1,271 rural Guatemalan children examined every three months up to age two and every six months thereafter as part of the INCAP study; 397 rural Bangladeshi children examined monthly to age one and quarterly thereafter as part of the Meheran Growth and Development Study; 468 rural Indonesian children examined monthly as part of the Ngaglik study; and 114 urban Japanese children examined monthly in studies from 1910 and 1920. Although all four studies were longitudinal, many observations from the Guatemala and Bangladesh studies were effectively cross-sectionally observed. Three different parametric forms were used to model the eruption process: a normal distribution, a lognormal distribution, and a lognormal distribution with age shifted to shortly after conception. All three distributions produced reliable estimates of central tendencies, but the shifted lognormal distribution produced the best overall estimates of shape (variance) parameters. Estimates of emergence were compared to other studies that used similar methods. Japanese children showed relatively fast emergence times for all teeth. Bangladeshi and Javanese children showed emergence times that were slower than are found in most previous studies. Estimates of agenesis were not significantly different from zero for most teeth. One or two central incisors showed significant agenesis that ranged from 0.1 to 0.8% in three of the samples; even so, failure to model the agenic proportion did not seriously bias the estimates.

Bangladesh↗

Comparison of four rapid D-Dimer tests for diagnosis of pulmonary embolism.

Aim of this study was to evaluate rapid D-Dimer tests for their utility in diagnosis of acute pulmonary embolism (PE). Tests were performed in 183 consecutive pats referred for lung scanning because of clinically suspected PE. According to lung scans and the clinical course of disease 19 pats were classified to have PE with high probability and 164 with low probability. An ELISA (Agen) was used as the D-Dimer reference, and results compared with those of a turbidimetric (Behring), an immunofiltration (Nycomed), latex plasma and whole blood agglutination test (both Agen). There was a poor correlation between the turbidimetric test and either the ELISA (R = 0.38) and immunofiltration test (R = 0.49). The correlation between the ELISA and immunofiltration test was better (R = 0.73). The qualitative latex and whole blood agglutination tests were better fitted to ELISA since positive and negative samples were overlapped only in their 1st and 9th percentiles of ELISA values. The whole blood agglutination test was positive at lower ELISA values than the latex test. The highest sensitivity test for PE was the immunofiltration test (95%) (500ng/mL cut-off), followed by the turbidimetric method (89%) (66ng/mL), the ELISA (89%) (300ng/mL), the whole blood test (88%) and the latex test (68%). Specificity was lowest for the immunofiltration test (33%), intermediate (57-65%) for the turbidimetric and whole blood agglutination tests, and highest for the ELISA and the most insensitive latex test (76/77%). The whole blood assay was found to be the fastest and most suitable for bed site testing but weak positives were difficult to read. The immunofiltration test required plasma preparation but allowed objective semiquantitation of results. The less rapid turbidimetric assay was fully quantitative and objective.

Adolescent↗

[Doping-related problems in clinical practice].

The complexity of the new antidoping regulations of the International Olympic Committee (IOC), the International Federations (IF) and the National Olympic Committees (NOC) rises a lot of problems in handling the prescriptions of medication in athletes in the daily practice. In addition, several countries have passed antidoping laws which makes the prescription and the delivery of doping agens illegal. This may have severe consequences for the prescribing doctors. It is therefore the goal of the article to inform the practitioning doctors about the new antidoping regulations and their impact on prescribing or delivering potential doping agens to athletes. It will focus on the new dopinglists, the different doping control systems, the problems with the pharmacological treatment of certain diseases as asthma and the necessary reports which have to be sent to the NOC's or the IF's after prescribing certain medications or methods.

Adrenergic beta-Agonists↗

A new method for measuring D-dimer using immunoturbidometry: a study of 255 patients with suspected pulmonary embolism and deep vein thrombosis.

D-Dimer testing has been suggested as a non-invasive method for the exclusion of pulmonary embolism (PE) and deep vein thrombosis (DVT). In this study, we compared a new method, the Miniquant D-dimer (Biopool International, Ventura, California, USA) to other previously validated D-dimer methods used for the purpose. Patients who were undergoing a definitive diagnostic study for thromboembolism had a blood sample drawn at that time. A whole-blood D-dimer (SimpliRed; Agen Biomedical Ltd, Brisbane, Australia) test was performed, and residual plasma was frozen and later analyzed using two enzyme-linked immunosorbent assay (ELISA) methods (D-dimer Gold; Agen, and Asserachrome D-Di; Stago International, Parsippany, New Jersey, USA) and the Miniquant D-dimer. Once all samples were analyzed, the correlation and accuracy of the Miniquant was compared with the ELISA method using Spearman's regression and Dunn's multiple paired comparison. All D-dimer methods were compared with radiographic studies. The data was analyzed collectively and segregated into in-patient (n = 112) and out-patient (n = 143) populations. The Miniquant D-dimer sensitivity, specificity and negative predictive value (NPV) for all patients were 95, 21, and 94% for DVT, and 100, 26, and 100% for PE. This new D-dimer method demonstrates acceptable sensitivity in patients with PE and DVT and, based on the high NPV of this method, it can be used for the exclusion of thromboembolism.

Adult↗

Comparison of six D-dimer methods in patients suspected of deep vein thrombosis.

We evaluated six D-dimer methods to determine their sensitivity, specificity, and negative predictive values (NPV) in symptomatic patients suspected of deep vein thrombosis (DVT). In patients suspected of DVT a whole blood D-dimer test (SimpliRED, Agen) was performed, and then tested using enzyme-linked immunosorbent assay (VIDAS D-Dimer, BioMerieux; Asserachrome D-Di, Stago International; Dimertest Gold, Agen) and automated immunoturbidometric methods (Advanced D-Dimer, Dade Behring; MiniQuant, Biopool). Each D-dimer method was independently compared with radiographic results to determine sensitivity and NPV. There were 151 patients enrolled in the study. Thirty-five (23.2%) patients had a positive Doppler ultrasound, with 26 proximal, eight distal, and one patient with both proximal and distal thrombus. Two patients (1.3%) had inconclusive studies and were excluded from the analyses. For all patients, the sensitivities for the rapid D-dimer methods were: SimpliRED, 82.3% [95% confidence interval (CI), 80.3-84.3%]; VIDAS D-Dimer, 91.4% (95% CI, 89.9-92.9%); MiniQuant D-Dimer, 96.3% (95% CI, 95.1-97.5%); and Advanced D-Dimer, 97.1% (95% CI, 96.3-97.9%). The sensitivity improved for SimpliRED (86.4%; 95% CI, 83.3-89.4%), VIDAS D-Dimer (95.5%; 95% CI, 85.0-100%), MiniQuant D-Dimer (100%; 95% CI, 96.9-100%) and Advanced D-Dimer (100%; 95% CI, 98.9-100%) in the inpatient population. The automated immunoturbidometric methods, the MiniQuant D-Dimer and Advanced D-Dimer, demonstrated comparable sensitivities and NPV with the VIDAS D-Dimer method in symptomatic patients suspected of DVT, which would suggest that these newer D-dimer methods could be used as part of the diagnostic algorithm for patients suspected of DVT.

Enzyme-Linked Immunosorbent Assay↗

Evaluation of a 2-minute anti-human immunodeficiency virus (HIV) test using the autologous erythrocyte agglutination technique with populations differing in HIV prevalence.

A total of 1,800 blood specimens (1,000 from healthy blood donors, 300 from patients with sexually transmitted disease, and 500 from intravenous drug users) were simultaneously tested with anti-human immunodeficiency virus enzyme-linked immunosorbent assay (ELISA) kits and a newly developed 2-min test for anti-human immunodeficiency virus based on the principle of autologous erythrocyte agglutination (AGEN Biomedical Limited). We found that AGEN's rapid test was as sensitive and specific as the other ELISA kits.

AIDS Serodiagnosis↗