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PubMedDate not supplied

Detection of recombinant epoetin and darbepoetin alpha after subcutaneous administration in the horse.

A direct detection method for anti-doping control of recombinant human erythropoietin (rHuEPO) abuse in racehorses is proposed. This method involves screening of plasma (or serum) by an enzyme-linked immunosorbent assay specific for human EPO and confirmation in urine samples by characterization of the urinary EPO isoelectric profile. This method was tested on horses that were administered epoetin alpha (rHuEPO) and the hyper-glycosylated form of this drug (darbepoetin alpha).

Animals↗
PubMed1993

Detection and quantitation of human immunodeficiency virus type 1 using molecular DNA/RNA technology.

The direct detection and quantitation of human immunodeficiency virus type 1 are becoming increasingly important for diagnosis and viral load determinations in human immunodeficiency virus type 1 infection. Traditional molecular-based assays such as Southern, Northern, and in situ hybridization have had limited application in this area due to the relatively low number of proviral copies per cell, the low percentage of infected cells, and transcriptional dormancy in many of the infected cells. Recent technological advances in polymerase chain reaction assays promise a number of useful applications in the clinical research and clinical laboratory setting.

Acquired Immunodeficiency Syndrome↗
PubMed1990

Organization and interaction of cholesterol and phosphatidylcholine in model bilayer membranes.

The molecular organization of sterols in liposomes of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) at 37 degrees C is examined by utilizing the fluorescent analogue of cholesterol cholesta-5,7,9-trien-3 beta-ol (cholestatrienol). (1) Cholestatrienol is shown to be indistinguishable from native cholesterol in terms of its ability to condense POPC, as determined by (i) pressure/area studies of mixed-lipid monolayers and (ii) its ability to increase the order of POPC bilayers (determined by electron spin resonance studies) whether on its own or admixed with cholesterol at various ratios. (2) By analysis of the perturbation of the absorption spectra, cholestatrienol was found to be freely miscible in aggregates of cholesterol in buffer. In contrast, a lack of any detectable direct interaction of the sterol molecules in POPC bilayers was detected. (3) Fluorescence intensity and lifetime measurements of POPC/sterol (1:1 mol/mol) at various cholesterol/cholestratrienol molar ratios (0.5:1 up to 1:1 cholestatrienol/POPC) confirmed that sterol molecules in the membrane matrix were not associated to any great degree. (4) A quantitative estimate of how close sterol molecules approach each other in the membrane matrix was evaluated from the concentration dependence of the steady-state depolarization of fluorescence and was found to be 10.6 A. From geometrical considerations, the sterol/phospholipid phase at 1:1 mol/mol is depicted as each sterol having four POPC molecules as nearest neighbors. We term this arrangement of the lipid matrix an "ordered bimolecular mesomorphic lattice". (5) The concentration dependence of depolarization of fluorescence of cholestatrienol in POPC liposomes in the absence of cholesterol yielded results that were consistent with the cholestatrienol molecules being homogeneously dispersed throughout the phospholipid phase at sterol/POPC ratios of less than 1:1. (6) From qualitative calculations of the van der Walls' hydrophobic interactions of the lipid species, the phospholipid condensing effect of cholesterol is postulated to arise from increased interpenetration of the flexible methylene segments of the acyl chains, as a direct result of their greater mutual attraction compared to their attraction for neighboring sterol molecules. (7) The interdependence of the ordered bimolecular mesomorphic lattice and the acyl chain condensation is discussed in an effort to understand the ability of cholesterol to modulate the physical and mechanical properties of biological membranes.

Buffers↗
PubMed1993

Rapid detection of Streptococcus pyogenes in pediatric patient specimens by DNA probe.

A chemiluminescent DNA probe test (Group A Streptococcus Direct Test; Gen-Probe, Inc., San Diego, Calif.) for rapid, direct detection of cRNA of Streptococcus pyogenes in throat swabs was compared with conventional culture and identification techniques. Throat swabs from 277 patients suspected of having streptococcal pharyngitis were examined. By DNA probe alone, 10 specimens were positive, 51 were positive by both assays, and 8 were positive by culture alone. Thus, DNA probe sensitivity, specificity, and positive and negative predictive values were 86, 95, 84, and 96%, respectively. Including an indeterminate category, sensitivity, specificity, and positive and negative predictive values were 89, 96, 86, and 97%, respectively. After discrepancy testing, these values for the raw data improved to 90, 98, 93, and 97%, respectively. None of the 24 specimens that grew non-S. pyogenes beta-hemolytic streptococci in culture were positive by the DNA probe. Because mucoid S. pyogenes strains are more virulent than nonmucoid strains, 24 isolates were retrospectively tested with the DNA probe to ensure that both types would be detected equally well. Isolates were examined in pure cultures as well as mixed with representative normal oral flora. There was no statistical difference in detection of any of the four groups. Group A Streptococcus Direct Test is a rapid, sensitive, and specific test for S. pyogenes.

Child↗
PubMed1997

Comparison of Technetium-99m sestamibi and indium-111 octreotide imaging in a patient with Ewing's sarcoma before and after stem cell transplantation.

BACKGROUND: We report the use of two novel nuclide agents, Technetium-99m (99Tc)sestamibi (MIBI) and indium-111 (In-111) octreotide, in comparison with conventional computed tomography (CT) imaging in a patient with metastatic Ewing's sarcoma (ES) before and after high dose chemotherapy with autologous peripheral stem cell transplantation (PSCT). MIBI is taken up actively by metabolically active tumor cells. Octreotide, a somatostatin analog, binds specifically to somatostatin receptors. METHODS: The patient was a 20-year-old male with recurrent metastatic ES to the lung. Before and sequentially after high dose chemotherapy and PSCT, the patient was imaged with MIBI. Whole body planar and single photon emission computed tomography (SPECT) images were obtained after the injection of 30 mCi of 99Tc MIBI. Prior to PSCT the patient was imaged with 6 mCi In-111 pentreotide. RESULTS: Conventional CT scans also were performed. Initial CT revealed pulmonary metastasis in the right lower lobe along with multiple left pleural-based lesions. These lesions were visualized clearly with MIBI. Octreotide detected only the left lung involvement. Sequential MIBI scans after PSCT correlated with tumor reduction in the right lung field and tumor progression in the left lung as well as the development of new pulmonary metastasis. These findings were confirmed on CT. CONCLUSIONS: MIBI imaging was highly concordant with CT scanning in the detection of metastatic ES. MIBI scanning holds promise for the direct detection of a variety of human malignancies, and may prove useful as a rapid whole body imaging modality.

Adult↗
PubMed2005

Further development of the beta-lactamase MutaGen assay and evaluation by comparison with Ames fluctuation tests and the umu test.

A rapid, high-throughput bacterial mutagenicity test system has been developed (MutaGen test) that detects reversions of inactivating base-pair substitutions and frameshifts in a TEM-1 class A beta-lactamase (ampicillinase) gene. To quickly and sensitively detect mutagens, the system utilises a series of plasmids that contain the mutated ampicillinase gene and the mucAB operon. Inactivating mutations in the ampicillinase gene include frameshifts integrated into repetitive GC-sequences and G-runs known to be mutagenic hot-spots, and base-pair substitutions inserted in or around the beta-lactamase active site. Frameshift mutations completely inactivated the enzyme only when located downstream of the active-site serine (Ser68). Previous (reporter gene based) assays with this system have detected reversion to ampicillin resistance by luminescence driven by induction of the tet-promotor controlled lacZ gene. In the present study, we describe the construction and evaluation of 19 additional potential tester strains. We also developed conditions for detecting reversions by pH shift using bromocresol purple and by directly detecting the enzymatic activity of beta-lactamase using nitrocefin. A 384-well format version of the pH shift MutaGen test was used to assay more than 20 chemicals. The responses in the assay were compared with responses for the same chemicals in the umu test and Ames fluctuation assays. The results indicate that the MutaGen test has high specificity for detecting specific mutations and, in some instances, better sensitivity than the other tests. Since the test is easy to conduct, sterile working conditions are not necessary, and the mutagenicity results are available either within one working day or overnight, the assay shows promise for the rapid screening of potentially genotoxic substances.

Bacteria↗
PubMed1994

Ionspray mass spectrometry of ciguatoxin-1, maitotoxin-2 and -3, and related marine polyether toxins.

A range of marine polyether toxins from dinoflagellates were analysed by ionspray mass spectrometry. Ciguatoxin-1 ([M+H]+ m/z = 1,111.8) purified from several fish species yielded singly charged ions corresponding to the parent ion, sodium and H2O adducts and ions for the loss of up to five H2O molecules. Ciguatoxin-1 was detected to 1 ng; however, interference from fish lipids precluded direct detection of ciguatoxin-1 in crude extracts from fish flesh spiked with ciguatoxin-1 at a level equivalent to 1.5 ng ciguatoxin-1/g of extracted flesh. Maitotoxin-2 yielded doubly and triply charged ions for sodium and potassium salts and likely possessed only one sulphate ester (M(r) = 3,298 for the mono-sodium salt). Maitotoxin-3, a recently isolated small maitotoxin, yielded singly charged ions including ions for the loss of one sulphate and up to four H2O molecules. Maitotoxin-3 is proposed to be a polyether compound possessing two sulphate esters (M(r) = 1,060.5 for the disodium salt). Brevetoxin-A ([M+H]+ m/z = 867.5) and brevetoxin-B ([M+H]+ m/z = 895.5) yielded singly charged ions corresponding to the parent ion, Na+ adducts and the loss of up to four H2O molecules. Okadaic acid ([M+H]+ m/z = 805.5) yielded singly charged ions corresponding to the parent ion and ions for the loss of up to three H2O molecules. A signal for M + 18 Da species that may represent [M+NH4]+ was observed for ciguatoxin-1, brevetoxin-A and -B, and okadaic acid. For all polyethers examined, the orifice potential influenced the relative intensity of the ions detected in a predictable manner.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1998

The use of genetic instability as a clinical tool for cancer diagnosis.

Human tumors exhibit two fundamentally important characteristics, extensive genetic alteration and clonality. Although it is still unclear to what extent tumors have an elevated mutational burden as compared with normal tissue, their clonality results in their ready detection. Thus, assaying tissues for clonal alterations at frequently mutated microsatellite loci represents a viable approach to cancer diagnosis. The most remarkable extension of this concept is that not only can cancer cells be detected in biological samples, but tumor DNA can also be directly detected in the serum or plasma of patients with some forms of cancer. This recent finding is currently being explored but may represent an important contribution to future diagnostic strategies.

DNA, Neoplasm↗
PubMed1992

Molecular detection of genetic defects in congenital adrenal hyperplasia due to 21-hydroxylase deficiency: a study of 27 families.

Congenital adrenal hyperplasia (CAH) due to 21-hydroxylase (21-OHase) deficiency is inherited as an autosomal recessive trait. Patients can present with the salt wasting, simple virilizing or a non-classical form of the disease. The gene for P450C21, the enzyme carrying 21-OHase activity, has been mapped to the major histocompatibility complex on chromosome 6p. Using molecular hybridisation techniques we have studied the genetic defect in 27 families with one or more affected offspring diagnosed and treated at the University Hospital of Essen. DNA samples were digested with restriction endonuclease TaqI, PvuII, BglII, and EcoRI and analysed by Southern blot hybridisation with the cDNA probe pC21/3c. Eleven of 40 haplotypes associated with the salt wasting form were found to have a large deletion of 30 kb affecting the 5' end of the active 21-OHase gene and the 3' end of the closely linked pseudogene. Results in another 11 cases are compatible with gene conversion; 18 cases were not informative. The 30 kb deletion was associated with a combination of the HLA antigens Bw47 and DR7 in 7 of 11 cases. In the haplotypes with gene conversion, no linkage disequilibrium to HLA antigens was found. No apparent gene alterations were detected in simple virilizing and non-classical haplotypes. The direct detection of the genetic defect in 55% of the salt wasting haplotypes may help to improve predictive testing in families with CAH.

Adrenal Hyperplasia, Congenital↗
PubMed1989

The incidence of legionella pneumonia: a 1-year prospective study in a large community hospital.

In a 1-year prospective study, 28 of 476 pneumonia patients (5.9%) were diagnosed as having legionella pneumonia. Legionella pneumonia was diagnosed in 12 (5%) of 240 community-acquired pneumonias and in 16 (6.8%) of 236 nosocomial pneumonias. Four methods of establishing diagnosis were used: serum antibody detection, direct proof of legionellae by immunofluorescence, culture, and legionella antigen detection in urine. The latter method proved to be clearly advantageous to all other methods in establishing the diagnosis. Twelve of the 28 patients (42.8%) died; all of them had severe underlying diseases. The fatality rate in patients treated with erythromycin was 18.8% (3/16 cases) and 75% (9/12 cases) in patients treated with antibiotics other than erythromycin. In 57.1% (16/28 patients) legionella pneumonia was acquired nosocomially. The results of our study underscore the need to use a broad spectrum of legionella diagnostic methods routinely and to administer antibiotics effective against legionellae in cases of pneumonia of unknown cause.

Adult↗
PubMed2004

Perception of non-voluntary brief contractions in normal subjects and in a deafferented patient.

The accuracy of force perception by human subjects in the absence of voluntary motor command was evaluated by exploring how they perceived isometric twitches of wrist extensor muscles produced by external stimulation. Twelve normal subjects and a well-known patient lacking large-diameter afferent fibres (GL) performed estimation, production and reproduction tasks. Magnetic stimulation of the radial nerve and, in normal subjects, transcranial magnetic stimulation (TMS) of the motor cortex were used to produce weak brief non-voluntary twitches. In estimation tasks, the subjects had to ascribe verbal marks on a 1-5 scale to the forces of stimulation-induced twitches. Loose covariations of marks and forces were observed, while directions of force variations between successive twitches were relatively well detected. GL did less well than normal subjects in detecting directions of force variations. In production tasks, subjects had to produce twitches matching verbal command marks in a 1-5 range, with or without visual feedback. Performances of normal subjects and GL resembled those of estimation tasks and were not improved by visual feedback. In reproduction tasks, subjects had to duplicate stimulation-induced test twitches: first without visual feedback, second with and third again without. Large errors were observed but all subjects did better with visual feedback. In the third step, improvement with respect to the first one was significantly more marked with TMS than with peripheral stimulation. GL improved her performance in the third step, possibly because she could use information provided by group III and group IV afferents still present in her nerves. Altogether, for normal subjects (1) the performances in estimation tasks are consistent with the known behaviour of Golgi tendon organs as observed in animal experiments, and (2) results observed in reproduction tasks suggest that cortical stimulation might elicit, in addition to corticospinal activation of motoneurones, collateral discharges that could be stored as a memory of motor command.

Adult↗
PubMed1990

Detection of C3bBb-stabilizing activity (C3 nephritic factor) in the serum from patients with membranoproliferative glomerulonephritis.

It is known that membranoproliferative glomerulonephritis (MPGN), hypocomplementaemia and C3 nephritic factor (C3NeF) are closely related to each other, and the presence or absence of C3NeF in the serum is important for evaluating the nature of MPGN. However, some difficulties have been encountered in detecting this factor and therefore a new assay permitting the direct detection of C3NeF without purifying IgG from the patient's serum has been devised. Using this assay method, C3bBb-stabilizing activity was observed even in sera from MPGN patients who were non-hypocomplementaemic. Furthermore, among 98 cases with hypocomplementaemia. C3NeF was found to be absent in 66 cases.

Complement C3 Convertase, Alternative Pathway↗
PubMed2002

Use of a nested polymerase chain reaction (N-PCR) to detect Trypanosoma cruzi in blood samples from chronic chagasic patients and patients with doubtful serologies.

Chagas disease, caused by Trypanosoma cruzi, is an important endemic illness in Latin America. Serologic tests for T. cruzi detection in blood are sensitive, but their specificity is unsatisfactory. Direct detection of parasites in blood, either by xenodiagnosis or hemoculture, is highly specific but of low sensitivity. Molecular assays such as the Polymerase chain reaction (PCR), which amplifies certain repetitive sequences of nuclear DNA has been used as a good alternative tool for T. cruzi detection in human blood. The present study aimed to test PCR diagnosis in chagasic chronic patients and doubtful serologic patients attended in GEDOCH (Chagas Disease Study Group/UNICAMP, Brazil). A 149 bp fragment originated from nuclear DNA was specifically detected in chronic chagasic patients. The results of these tests were compared with serologic diagnosis performed using standard techniques and xenodiagnosis. We found that 43 out of 50 patients previously serodiagnosed as chagasic were positive using the N-PCR method. Thirteen of 30 patients with doubtful serologic results were confirmed as positive by N-PCR. Our results suggest that the N-PCR may be a complementary tool to serology in the diagnosis of Chagas disease, and that it is usefull for parasite detection in patients with chronic disease and patients with doubtful serologic results.

Animals↗
PubMed2000

Surface-enhanced raman detection of 2,4-dinitrotoluene impurity vapor as a marker to locate landmines.

Time, cost, and casualties associated with demining efforts underscore the need for improved detection techniques. Reduction in the number of false positives by directly detecting the explosive material, rather than casing material, is desirable. The desired field sensor must, at a minimum, demonstrate reproducibility, the necessary level of sensitivity, portability, instrumental stability, and fast system response times. Ideally, vibrational spectroscopic techniques have the potential to remove false positives, since every chemical has a unique bond structure. Herein, we demonstrate the capabilities of surface-enhanced Raman spectroscopy to detect the chemical vapor signature emanating from buried TNT-based landmines. We present reproducible results obtained from blind tests controlled by the Defense Advanced Research Projects Agency (DARPA) that demonstrate vapor detection of 2,4-dinitrotoluene at concentration levels of 5 ppb or less. The results presented used acquisition times of 30 s on a fieldable system and showed that SERS can be a significant improvement over current landmine detection methods.

Journal Article↗
PubMed2002

Evaluation of an amplified Mycobacterium tuberculosis direct test in clinical specimens.

SETTING: An increase in tuberculosis cases has been observed since the last decade, and rapid and specific tests are needed to directly detect Mycobacterium tuberculosis in respiratory specimens. The Amplified Mycobacterium Tuberculosis Direct Test (AMTDT) is a direct specimen assay for the identification of M. tuberculosis. OBJECTIVE: To evaluate the sensitivity and specificity of the AMTDT. METHODS: We performed the test in 426 patients: 58 patients who had culture-proven and/or clinically diagnosed tuberculosis, and 368 patients who were negative for M. tuberculosis by culture and clinical criteria (the control group). The results were compared to culture and clinical diagnosis of tuberculosis. RESULTS: AMTDT was found to be positive in 35 of the 58 patients who had culture-proven and/or clinically diagnosed tuberculosis and 71 of the 368 controls. Seventeen of the tuberculosis group were smear-negative, and AMTDT was positive in 11 of these patients. The sensitivity and specificity of the test were respectively 60% and 80%. CONCLUSION: AMTDT can be used as a complementary technique in tuberculosis patients in the presence of high clinical suspicion.

Diagnosis, Differential↗
PubMed1996

[Respiratory syncytial virus lower respiratory tract infection: our series].

The authors report 57 cases of Respiratory Syncytial Virus (RSV) lower respiratory tract infection. The diagnosis was performed by detection of specific serum antibodies of IgM class or by direct detection of RSV antigens in nasopharyngeal specimens. Most of the patients were younger than one year. No one had risk factors for severe RSV infection form. All patients recovered. The authors emphasize the difficulties to discriminate between bronchiolitis and interstitial pneumonitis on the basis of the clinical and radiological features.

English Abstract↗
PubMed2006

[West Nile virus fever].

West Nile virus (WNV) is a member of the Flaviviridae family, genus Flavivirus. Its reservoir hosts are wild birds. Infection is transmitted to humans by infected mosquitoes of the genus Culex. In most cases, it is either asymptomatic or manifests itself as mild fever. Typically, WNV illnesshas a sudden onset with fever above 39 degrees C and accompanying symptoms such as chills, headache, arthralgia, myalgia, back ache, cough and sore throat. Gastrointestinal symptoms are frequently reported. Generalized lymphadenopathy and conjunctivitis may develop. In some patients the infection can progress to meningoencephalitis. Diagnosis is currently based on detection of IgM antibodies in blood and cerebrospinal fluid or direct detection of WNV RNA.

Humans↗
PubMed1997

Detection methods of Helicobacter pylori: accuracy and costs.

A variety of methods exist for determining gastric colonization with Helicobacter pylori, which has been implicated in the development of peptic ulcer disease. The goal of this study was to evaluate four of the current methods available in a clinical surgical practice setting through a prospective evaluation of 40 consecutive patients undergoing upper diagnostic endoscopy. All patients underwent six antral gastric biopsies for use with the following detection methods: histologic demonstration of organisms (hematoxylin and eosin stain), direct detection of urease activity (Remel Selective Rapid Urea, Lenexa, KS), and culture of H. pylori. All patients also had measurement of serum immunoglobulin G for H. pylori by the enzyme-linked immunosorbent assay method (Corning Clinical Laboratories, St. Louis, MO). The infection status was established by a concordance of test results. The results show that H. pylori can be assessed equally well with histology, a rapid urease test, and serology, with all three tests having good sensitivity (92-100%) and specificity (85-96%). The culturing of the organism had poor sensitivity (42%). The benefits of the urease test are a much more rapid response time and a much lower cost as compared to histologic and serologic testing. In conclusion, the rapid urease test is the method of choice to detect H. pylori in those patients undergoing endoscopy in whom the identification of H. pylori will change their management.

Adolescent↗