Prenatal diagnosis of compound heterozygous deficiency of protein C by direct detection of the mutation sites.
Explore the source record for details and available documents.
Genetic Testing↗
SEARCH · Search PubMed
Results for “Direct detection”
Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.
Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.
Direct detection of multiple point mutations in mitochondrial DNA.
Mitochondrial defects can be caused by mutations in nuclear or mitochondrial DNA. Large deletion/duplication and point mutations are the two major types of mitochondrial DNA (mtDNA) mutations. Comprehensive molecular diagnosis requires the analysis of multiple point mutations. We developed an effective multiplex PCR/allele-specific oligonucleotide (ASO) method to simultaneously screen multiple point mutations in mtDNA. The system involved three pairs of primers to amplify mutation "hot spots" at tRNA(leu(UUR)), tRNA(lys)/ATPase, and ND4 regions, followed by detection of point mutations with ASO probes. Over 2000 specimens were analyzed and the results were compared with those from previous studies with the PCR/restriction fragment length polymorphism method. Our data demonstrate that the multiplex PCR/ASO method is much more sensitive in the detection of low mutant heteroplasmy. It is simple and cost effective, especially if a large number of samples are to be screened for multiple point mutations.
DNA, Mitochondrial↗
Direct detection of premature protease activation in living pancreatic acinar cells.
Explore the source record for details and available documents.
Animals↗
[Improvement and acceleration of the diagnosis of contagious bovine pleuropneumonia by direct detection of the microbe using polymerase chain reaction (PCR)].
A diagnostic procedure is described for the detection of Mycoplasma mycoides subsp. mycoides SC, the causative agent of contagious bovine pleuropneumonia. DNA extracted from clinical samples was investigated by the polymerase chain reaction (PCR) using at least 2 different primer pairs, one species-specific and another one specific for the class of Mollicutes. Using this method, the time required for detection of the pathogen was reduced to 2 days, whereas with traditional diagnostic methods (cultivation in broth, biochemical tests or immunofluorescence) the same finding would be available only within approximately 20 days. Although contagious bovine pleuropneumonia does not occur in Central Europe, there are occasional identifications of cattle having positive titres in the complement fixation test (CFT). Immunoblotting analysis of such sera confirmed that the reason for this phenomenon were cross-reactions with taxonomically related mycoplasma species. The present PCR assay proved to be suitable because of its rapidity, as well as high specificity and sensitivity. In the case of positive serological findings it enables diagnosticians to provide evidence on the presence or absence of the agent at short notice.
Animals↗
Rapid viral diagnostic techniques.
The application of viral diagnosis to clinical medicine is approaching a critical period in which greater emphasis will be placed on altering patient care as a result of viral identification. Although isolation of viruses in tissue culture is the "gold standard" to which all other methods of viral identification are compared, the expense in terms of cost and technical time required as well as the time necessary for specific viral isolation has severely limited the use of viral diagnosis in clinical medicine. The concomitant development of rapid viral diagnostic procedures with the increased availability of anti-viral therapy, promises to alter how patients will be managed in the future. The greatest emphasis for rapid viral diagnosis has been placed on viral detection directly from clinical specimens since these procedures eliminate the need to cultivate the virus. Methods for direct detection of viral antigens in clinical specimens include electron microscopy and a wide range of immunologic assays. The extraction of nucleic acid directly from clinical specimens and hybridization of this DNA to specific viral probes also provides for virus identification without the need to culture the virus. Currently, many of the rapid diagnostic procedures discussed are performed only in research laboratories. Over the next several years, however, methods of antigen detection using EIA procedures and detection of viral-specific nucleic acid promise to alter how we study and diagnose viral infections.
Humans↗
Direct characterization of Blastocystis from faeces by PCR and evidence of zoonotic potential.
In vitro propagation followed by PCR, and a PCR-based method capable of the direct detection of Blastocystis in faeces were utilized to detect Blastocystis from various hosts in Australia, including primates and their handlers from the Perth Zoo. In addition, Blastocystis isolates from dogs and humans living in a localized endemic community in Thailand were also characterized genetically. PCR-based detection directly from faeces was shown to be more sensitive compared with in vitro culture for the detection of Blastocystis. Moreover, phylogenetic analysis of Blastocystis isolates amplified utilizing in vitro techniques prior to PCR revealed that this method favoured the preferential amplification of Blastocystis subtype 5 over subtype 1. This study is the first to provide molecular-based evidence supporting the zoonotic potential of Blastocystis in dogs, possums and primates in a natural setting.
Animals↗
Use of Dimethyl Sulfoxide to Detect Hydroxyl Radical during Bacteria-Induced Hypersensitive Reaction.
Excess active oxygen is generated during the hypersensitive reaction (HR), an incompatible reaction of plants to bacterial pathogens. During HR, lipid peroxidation correlates chronologically with production of the oxygen species, superoxide (O(2) (.-)). However, O(2) (.-) may not be the active oxygen species that initiates lipid peroxidation. Evidence from other systems suggest that O(2) (.-) is converted to the hydroxyl radical (HO(.)) before lipid peroxidation is initiated. Until recently, HO(.) could not be detected directly in vivo. This study utilizes a newly reported method to directly detect and quantify the formation of HO(.)in vivo. Dimethyl sulfoxide (DMSO), used as a molecular probe, is oxidized by HO(.), forming the stable compound methanesulfinic acid. The methanesulfinic acid can be easily extracted from plant tissues and measured with a colorimetric assay. This study demonstrates significant increases in HO(.) concentration after simultaneous infiltration of cucumber (Cucumis sativa L.) plants with paraquat and DMSO. The concentration of HO(.) did not increase significantly when cucumber plants were infiltrated simultaneously with the HR-inducing bacteria, Pseudomonas syringae pv. pisi, and with DMSO. Lipid peroxidation, however, could be measured at times when HO(.) was not detectable. It appears that HO(.) is not generated during bacteria-induced HR; therefore, HO(.) is not responsible for the initiation of lipid peroxidation.
Journal Article↗
Spectroscopic studies of cutaneous photosensitizing agents. XVII. Benzanthrone.
The photochemistry of benzanthrone (7H-benz[de]-anthracene-7-one) has been studied using electron paramagnetic resonance (EPR) in conjunction with the spin trapping technique and the direct detection of singlet molecular oxygen luminescence. Irradiation (lambda ex = 394 nm) of benzanthrone (BA) in aerated ethanol, dimethylsulfoxide or benzene resulted in the generation of superoxide (O2-.) which was trapped by 5,5-dimethyl-1-pyrroline-N-oxide. The ethoxy radical was also detected in ethanol. Photolysis of BA in deaerated basic ethanol led to the formation of BA anion radical, BA-., which was detected directly by ESR. This radical anion decayed back to BA with a unimolecular rate constant of 1.5 x 10(-3) s-1. The 1O2 quantum yields (lambda ex greater than 345 nm) for BA in ethanol, 90% ethanol and basic ethanol (0.1N NaOH) were 0.89, 0.88 and 0.28 respectively relative to Rose Bengal. The lower yield of 1O2 in basic ethanol may be attributable to the reaction of oxygen with BA-. (which is generated in higher yield at alkaline pH) to give O2-.. These findings suggest that on exposure to light BA can generate active oxygen species which may be responsible for the photocontact dermatitis caused by BA in industrial workers exposed to this chemical.
Animals↗
Comparison between electrochemical and optoelectrochemical impedance measurements for detection of DNA hybridization.
The principles of the electrochemical and optoelectrochemical impedance measurements on bare electrolyte/dielectric/semiconductor structures are described. The analysis of the experimental curves allows access to several indications concerning the electrical behavior of such structures. The application of these techniques to follow the electrical behavior of structures modified with two biological systems was investigated. The antibody/antigen recognition did not change the surface charge and, therefore, did not affect the impedance curves with respect to the applied potential. By contrast, the hybridization of two complementary DNA strands on the surface of the structure induced a variation of flat band potential of the semiconductor leading to a shift of impedance curves along the potential axis. This means that it is possible to detect directly the DNA hybridization without the use of labeled probes. The use of light allows the surface to be probed locally. In the future, the application of this technique for direct detection of hybridization on DNA chips should be possible.
Biosensing Techniques↗
Direct mecA detection from blood culture bottles by branched-DNA signal amplification.
A branched-DNA (bDNA) signal amplification method was used to detect the mecA gene directly from blood culture broth growing staphylococci. BACTEC blood culture bottles with positive growth indices and containing staphylococcus-like organisms as shown by Gram stain were tested for the presence of the mecA gene. Comparison of test results was done among 225 patients (one blood culture from each patient). Compared with PCR, the sensitivity and specificity of the bDNA method are 100 and 99%, respectively. The bDNA test is carried out in a 96-well format and requires approximately 6 h to perform. Our preliminary results suggest that direct detection of the mecA gene by bDNA signal amplification is (i) sensitive enough to detect mecA directly from blood culture bottles without the requirement for subculture and (ii) as sensitive and specific as the PCR-based method.
Bacteremia↗
A simple method to detect Plasmodium falciparum directly from blood samples using the polymerase chain reaction.
We have developed a simple method for treating blood samples permitting direct detection of Plasmodium falciparum parasites using the P. falciparum-specific DNA probe pPF14 after polymerase chain reaction (PCR) amplification of target DNA sequences, and have compared this method with microscopic examination of thick blood smears. For PCR amplification, blood samples were lysed, then filtered onto filter paper. After drying, a piece of the filter paper was added directly to the PCR mixture for amplification. The presence of PCR products was detected using nonisotopically labeled probe. This method permits detection of less than than 10 parasites in a 20-microliters sample, and minimizes the effects of PCR inhibitors generally found in blood. Samples were collected from patients presenting at malaria clinics in Mae Sod and Mae Ramat, Thailand, and 626 were analyzed both by the PCR method and by conventional microscopy. Of these, 157 were positive both by microscopy and by PCR, while 297 were negative by both methods. PCR detected 131 samples that were negative by microscopy, and failed to detect 41 samples identified as positive by microscopy. All discordant samples were re-analyzed by microscopy and by PCR. Upon re-examination at a higher sensitivity, microscopy identified five additional positive cases, while six previously positive cases were found to be negative. This method of treating blood samples for PCR may also be useful in other diagnostic assays.
Animals↗
Direct carbon detection in paramagnetic metalloproteins to further exploit pseudocontact shift restraints.
Direct detection heteronuclear NMR allows us to drastically reduce paramagnetic contributions to the line width as compared to 1H detection. As an example, a calcium binding protein (human oncomodulin), in which one of the calcium ions was selectively substituted with Tb3+, is used. Through a variety of 13C direct detection NMR experiments, resonances were detected as close as 5.5 A from the metal ion. Pseudocontact shifts measured through 13C direct detection experiments provide structural constraints in regions of the protein where 1H resonances are broadened beyond detection through Curie relaxation (up to 16 A from the paramagnetic center).
Calcium↗
Direct EPR detection of transient and continuous wave signals at 2.5 GHz.
Direct detection of free induction decays and electron spin echoes, and the recording of echo-detected EPR spectra and electron spin echo envelope modulation patterns at a microwave frequency of 2.5 GHz is demonstrated. This corresponds to the measurement of the transverse magnetization in the laboratory frame, rather than in the rotating frame as usually done by down-converting the signal (homodyne detection). An oscilloscope with a 6-GHz analog bandwidth, a sampling rate of 20 GigaSamples per second, and a trigger frequency of 5 GHz for the edge trigger and 750 MHz for the advanced trigger, is used in these experiments. For signal averaging a 3-GHz microwave clock divider has been developed to synchronize the oscilloscope with the frequency of the EPR signal. Moreover, direct detection of continuous wave EPR signals at 2.5 GHz is described.
Journal Article↗
Direct ESR detection of a free radical intermediate during the peroxidase-catalyzed oxidation of the antimalarial drug primaquine.
Oxidation of the antimalarial primaquine by horseradish peroxidase and H2O2 was demonstrated by visible light absorption and ESR spectroscopy. Initial product analysis indicated a 15% yield of O-demethoxylation products, methanol and the quinone-imine derivative, and organic extractable polymeric material. Horseradish peroxidase was substituted by methemoglobin, and both enzymes showed greater activity at acidic pH values. During the enzymatic oxidation of primaquine, a drug-derived free radical was detected by direct ESR spectroscopy. A similar ESR spectrum was obtained during enzymatic oxidation of 6-hydroxyprimaquine at pH 9.0. Computer simulations of the ESR spectra obtained in normal and deuterated buffer indicated that the detectable free radical contains two primaquine moieties. This in vitro oxidation of primaquine to a free radical intermediate that is stable in the presence of oxygen might be considered a new mechanistic route for analyzing the pharmacological effects of primaquine.
Electron Spin Resonance Spectroscopy↗
Direct optical detection of protein-ligand interactions.
Direct optical detection provides an excellent means to investigate interactions of molecules in biological systems. The dynamic equilibria inherent to these systems can be described in greater detail by recording the kinetics of a biomolecular interaction. Optical biosensors allow direct detection of interaction patterns without the need for labeling. An overview covering several commercially available biosensors is given, with a focus on instruments based on surface plasmon resonance (SPR) and reflectometric interference spectroscopy (RIFS). Potential assay formats and experimental design, appropriate controls, and calibration procedures, especially when handling low molecular weight substances, are discussed. The single steps of an interaction analysis combined with practical tips for evaluation, data processing, and interpretation of kinetic data are described in detail. In a practical example, a step-by-step procedure for the analysis of a low molecular weight compound interaction with serum protein, determined on a commercial SPR sensor, is presented.
Binding, Competitive↗
Measurement of phenols on a loop-supported liquid film by micellar electrokinetic chromatography and direct UV detection.
This paper describes a direct measurement of phenolic substances in the gas phase at levels relevant to occupational health by micellar electrokinetic chromatography (MEKC) using direct ultraviolet detection. A small circular loop (2 mm O.D.) is formed with 100 mm Pt-wire at the sampling end of a fused-silica capillary. The capillary tip of the sampling end is present at the center of the loop and in the same plane. A thin film of 0.50 mM NaOH is formed on the loop by immersion and withdrawal. The film is in fluid communication with the capillary and acts as an absorber for gaseous phenols. Gas sampling is performed by transferring the film-bearing loop into a chamber through which air is aspirated for a preset time at a preselected flow-rate (typically 1 min at 100 cm3/min). A part of the film content is then introduced into the capillary by gravity injection and then MEKC is performed. A total of twelve chloro- and nitrophenols were selected for this study. Under the above sampling conditions, limits of detection (LODs) for various phenols range from high-single-digit to low-double-digit ppbv levels. The effect of several critical parameters, such as the composition of the liquid film and the MEKC running electrolyte, sampling period and film evaporation on the collection efficiency, separation efficiency and calibration behaviour are described.
Adsorption↗
Direct MRI detection of neuronal magnetic fields in the brain: theoretical modeling.
Whether MRI can be used for direct detection of neuronal activity is a matter of debate. Controversial theoretical and experimental results have been reported. Here, we present an improved current-dipole model to compute magnetic field generated by neural firing and to calculate MRI signal changes resulting from the neuronal magnetic field (NMF). Each dendrite or each unmyelinated axon was modeled as a modified current-dipole. NMF were estimated based on a synchronized activity of multiple neurons. Sensitivity of using phase and magnitude MRI to measure effects of NMF was evaluated. Our results show that NMF can potentially generate up to a few percent changes in MRI magnitude signals. Phases of MRI signal tend to be destructively added and are insensitive to NMF in the activated region when the distribution of the activated dendrites is symmetrical. Phases could be detected when the distribution of the activated dendrites is asymmetrical and on some neighboring voxels. Our modeling implies that direct MRI detection of neuronal activity is possible.
Brain↗
Direct carrier detection in hemophilia B kindreds: use of modified primers (mutagenic primers) for enzymatic amplification of the factor IX gene.
Rapid direct detection of point mutations in hemophilia B kindreds was performed by analyzing the restriction fragments of the factor IX gene amplified by polymerase chain reaction (PCR). The family members of the two patients, HB 5 and HB 6, whose mutant factor IX gene had previously been defined by sequence analysis, were examined in this study. Since there are no restriction endonucleases available for detecting each mutation directly, we designed modified primers which were substituted one nucleotide near the mutated positions. Following PCR with these primers, new Mbo I or Alu I cleavage sites for normal alleles were created. In each family, the restriction fragment revealed the carriership of each patient's mother. This simple methodology to detect mutations of interest is useful for genetic counseling in sporadic cases of hemophilia B.
Alleles↗