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At least 235 records · Page 13Linked to original sources

PubMed2002

Fast analysis of flavonoids in plant extracts by liquid chromatography-ultraviolet absorbance detection on poly(carboxylic acid)-coated silica and electrospray ionization tandem mass spectrometric detection.

A highly hydrophilic poly(7-oxonorbornene-5,6-dicarboxylic acid-block-norbornene) [=poly-(ONDCA-b-NBE)]-coated silica was investigated for the liquid chromatographic (LC) determination of flavonoids in plant extracts of complex biological origin using UV absorbance and mass spectrometric (MS) detection. Compared to the most commonly used octadecyl derivatized silica this sorbent allowed fast separations even at extreme pH values. Furthermore, UV absorbance and MS detection were evaluated. As we found, UV detection at 254 nm allows the determination of flavonoids down to the ng range with a linearity of R2>0.9906. For the more selective characterization the validated LC system was coupled to a quadrupole ion trap mass spectrometer via an electrospray ionization (ESI) interface. MS detection showed high linearity (R2>0.9904) for all investigated flavonoids. Due to the relatively high flow-rate of 1 ml/min the limits of detection were found in the lower-microg range. Collision induced dissociation was applied to obtain characteristic fragmentation fingerprints. Finally, the validated LC-ESI-MS-MS method demonstrated that this poly-(ONDCA-b-NBE) stationary phase allows fast characterization and quantitation in onion, elderflower blossom, lime blossom, St. John's Wort and red wine.

Calibration↗

PubMed1985

Expert system for pharmaceutical analysis. I. Selection of the detection system in high-performance liquid chromatographic analysis: UV versus amperometric detection.

The usefulness of amperometric detection in pharmaceutical analyses was investigated for different groups of drugs. The UV response at 254 nm and that at the absorption maximum of the solute were compared with the electrochemical signal obtained. The minimum detectable concentration (nanograms on-column) of each substance is reported for the three different detection systems. This comparison was performed for 72 drugs (local anaesthetics, antipyretics, tricyclic antidepressants, sulphonamides, sex hormones, beta-adrenoceptor blocking agents, phenothiazines, alkaloids, diuretics and penicillins). The median limit of detection of the amperometric detector (see definition in the text) is 1.0 ng on-column and the median gain in sensitivity, compared with UV detection is 22.5.

Adrenergic beta-Antagonists↗

PubMed2002

Determination of pesticide residues in coconut water by liquid-liquid extraction and gas chromatography with electron-capture plus thermionic specific detection and solid-phase extraction and high-performance liquid chromatography with ultraviolet detection.

Two simple methods were developed to determine 11 pesticides in coconut water, a natural isotonic drink rich in salts, sugars and vitamins consumed by the people and athletes. The first procedure involves solid-phase extraction using Sep-Pak Vac C18 disposable cartridges with methanol for elution. Isocratic analysis was carried out by means of high-performance liquid chromatography with ultraviolet detection at 254 nm to analyse captan, chlorothalonil, carbendazim, lufenuron and diafenthiuron. The other procedure is based on liquid-liquid extraction with hexane-dichloromethane (1:1, v/v), followed by gas chromatographic analysis with effluent splitting to electron-capture detection for determination of endosulfan, captan, tetradifon and trichlorfon and thermionic specific detection for determination of malathion, parathion-methyl and monocrotophos. The methods were validated with fortified samples at different concentration levels (0.01-12.0 mg/kg). Average recoveries ranged from 75 to 104% with relative standard deviations between 1.4 and 11.5%. Each recovery analysis was repeated at least five times. Limits of detection ranged from 0.002 to 2.0 mg/kg. The analytical procedures were applied to 15 samples and no detectable amounts of the pesticides were found in any samples under the conditions described.

Chromatography, Gas↗

PubMed2003

Handling and detection of 25 amol of near-infrared dye deoxynucleotide conjugates by capillary electrophoresis with laser-induced fluorescence detection.

Near-infrared dyes are attractive as labeling reagents to enhance sensitivity in trace analysis largely because background fluorescence is low in this spectral region. Here we demonstrate, towards a goal of detecting DNA adducts in small biological samples, that some near-infrared (IR) dye-labeled deoxynucleotides can be separated and detected with high sensitivity by capillary electrophoresis (CE)-laser-induced fluorescence detection (LIF) in a realistic way (handling detection limit of 25 amol) for near-IR dye-labeled deoxynucleotides. This detection limit is achieved by polarity-switching injection of 2.0 microl from a volume of 5.0 microl, in which the compounds are 5 x 10(-12) mol/l in 50% aqueous methanol. Although the adenine and cytosine-containing conjugates co-migrated, the other three (guanine, N2-ethylguanine and thymine) were resolved.

Coloring Agents↗

PubMed1999

Enzyme amplification as detection tool in continuous-flow systems. II. On-line coupling of liquid chromatography to enzyme-amplified biochemical detection after pre-column derivatization with biotin.

Enzyme-amplified biochemical detection (EA-BCD) was used as a post-column detection technique, coupled on-line with high-performance liquid chromatography (HPLC). The enzyme detection system was developed to detect biotin or biotin containing compounds. Biotinylation is widely used to label analytes of interest ranging from small molecules to proteins and DNA. Naphthalene aldehyde and anthracene aldehyde were used as model compounds. Both compounds were biotinylated off-line with biotin aminocaproic hydrazide (BACH). On-column biotinylation was performed by preconcentration of anthracene aldehyde on copper phthalocyanine. After biotinylation, samples were introduced to the HPLC system. Enzyme-labeled streptavidin, which possesses high affinity to biotin, was added post-column to the HPLC effluent. Excess of enzyme-labeled affinity protein was removed by means of an immobilized biotin column. After separation of free and bound fraction, substrate was added, which was converted to a fluorescent product by the enzyme label. Using alkaline phosphatase as an enzyme label, a mass detection limit after on-column preconcentration and biotinylation of 250 fmol was achieved.

Biotin↗

PubMed1994

Characteristics of prostate cancer detected in the American Cancer Society-National Prostate Cancer Detection Project.

The American Cancer Society-National Prostate Cancer Detection Project is a prospective, comparative study of a cohort of 2,999 men 55 to 70 years old not suspected on entry of having prostate cancer. A total of 164 prostate cancers is available from this project for analysis. A small proportion of tumors detected were advanced in terms of the clinical stage at diagnosis. Cancer detected by digital rectal examination tended to be more advanced than that found on the basis of only transrectal ultrasound or prostate specific antigen (PSA). A large proportion of patients received curative therapy involving radical prostatectomy in 67.1% and radiotherapy in 18.3%. Of 103 men presumed to have organ confined disease and treated by prostatectomy 64 (37.9%) actually had locally extensive cancer pathologically. PSA level and PSA density were associated with the detection of organ confined cancer but several advanced tumors had PSA levels in the normal range. Age referenced PSA, compared to conventional standards, demonstrated lower sensitivity to cancer with little improvement in specificity. The disease resulting from this multimodality detection effort represented a spectrum of pathological conditions. Further followup and evaluation are needed to determine whether these benefits are reflected in long-term mortality and survival experience.

Aged↗

PubMed1999

Simple method of detecting enteroviruses in contaminated molluscs and sewage by using polymerase chain reaction coupled with a colorimetric microwell detection assay.

The methods normally used for the detection of enteroviruses in environmental samples involve the use of cell cultures, which are expensive and time consuming. The Polymerase Chain Reaction (PCR) is a useful tool for the detection of enteroviruses in several matrixes because primary cell culture is not needed and the increased sensitivity of PCR allows detection of the low numbers of target nucleic acids usually found in environmental samples. A 5-h, user-friendly PCR assay was used to detect enteroviruses in bivalves molluscs (clams) and sewage. Reverse transcription and amplification were performed in a one-step reaction using rTth polymerase. Carryover contamination was prevented with dUTP and uracil N-glycosylase. Detection was performed colorimetrically in a microwell titer plate. This method has greater advantages over conventional methodologies for routinely screening a large number of samples, namely, the rapid acquisition of results and cost effectiveness.

Animals↗

PubMed2003

Detection of petunia vein-clearing virus: model for the detection of DNA viruses in plants with homologous endogenous pararetrovirus sequences.

A number of cases of plant virus sequence integration into host plant genome have been reported. In at least two cases, endogenous pararetrovirus sequences are correlated strongly with subsequent episomal virus infection and there is circumstantial evidence that this also occurs for Petunia vein-clearing virus (PVCV). The detection of viruses is a critical component of plant health and therefore, it is important to have diagnostic procedures that differentiate between the detection of encapsidated viral DNA and homologous sequences in the host genome. PCR-based detection methods targeted at PVCV DNA have been tested and particular attention was paid to design controls that would indicate the existence of host DNA in the reaction. The use of ion-exchange chromatography for the partial purification of plant viruses from other cellular components, including chromosomal DNA, is described. The methods tested for PVCV detection are used to illustrate general principles for the specific detection of virus infections in host plants that carry homologous virus sequences in their genomes.

Chromatography, Ion Exchange↗

PubMed1997

Detection of tick-borne encephalitis virus by sample transfer, plaque assay and strand-specific reverse transcriptase polymerase chain reaction: what do we detect?

Experimental inoculation of mice provides a well characterized model for studying infection with tick-borne encephalitis virus (TBEV), a flavivirus pathogenic for humans. Conflicting data on the kinetics of viremia and the development of virus titers in the brain, however, were only recently shown to have resulted from the use of assay systems with different levels of sensitivity in the titration of TBEV, i.e. plaque assay or sample transfer into naive recipient mice. Theoretically, RT-PCR could extend further the detectability to antibody-neutralized virus and when undertaken strand-specifically discriminate active replication from the mere presence of TBEV. We have compared the conventional methods for detection of TBEV with a newly devised RT-PCR method. As expected, RT-PCR, in contrast to the infectivity assays, detected antibody-neutralized virus. Furthermore, the mere presence or active replication of the virus could be differentiated by strand-specific RT-PCR. Plaque assay and sample transfer, in contrast, both detected only infectious virus. However, whereas sample transfer provides higher sensitivity for detection of TBEV from solid organs, the plaque assay is less costly and considering animals welfare more convenient. Thus, the newly devised method may allow the resolution of unanswered questions, while both the traditional infectivity assays retain their benefits in certain situations.

Animals↗

PubMed1999

Highly sensitive detection of swine vesicular disease virus based on a single tube RT-PCR system and DIG-ELISA detection.

A highly sensitive detection of swine vesicular disease virus (SVDV) based on a single tube RT-PCR system and digoxigenin (DIG)-PCR-ELISA detection was developed. Using a one tube RT-PCR system, optimisation of the PCR conditions and optimisation of the microwell hybridisation and colourimetric detection of the amplicons resulted in a method that could detect viral RNA in infected tissue culture fluid with a titre as low as 0.1 TCID50/100 microl. The same sensitivity was obtained with SVDV-spiked faeces, if the samples were pre-treated with 1,1,2-trichlorotrifluoroethane/chloroform and subsequently concentrated using an ultrafiltration system and RNA extracted with the Purescript kit. The specificity of the test was validated on 27 SVDV strains belonging to four different groups. No cross-reactivity with genetically and symptomatically related viruses was detected using RNA of foot-and-mouth disease virus (FMDV), porcine enterovirus (PEV), vesicular stomatitis virus (VSV), Coxsackie B5 virus (CV-B5) and encephalomyocarditis virus (EMCV). The test was validated successfully on clinical samples, being slightly more sensitive and much faster than virus isolation on cell cultures. Moreover the possibility of automating the procedure will allow the processing of large numbers of clinical samples.

Animals↗

PubMed2004

How useful is glucose detection in diagnosing cerebrospinal fluid leak? The rational use of CT and Beta-2 transferrin assay in detection of cerebrospinal fluid fistula.

BACKGROUND: This report describes the sensitivity and specificity of glucose detection using Glucostix test strips and computed tomography (CT) of the skull base for confirming cerebrospinal fluid (CSF) fistulae in patients with persistent rhinorrhoea or otorrhoea, and comparing them with the beta-2 transferrin assay as the gold standard for CSF detection. METHODS: Fluid samples from the nose were collected from 18 patients with suspected CSF fistulae. The samples were assayed for beta-2 transferrin using the Western blotting and immunostaining technique. CT (5mm axial slice) of the skull base was performed for evidence of skull base fracture. The glucose levels and Glucostix results were compared. RESULTS: Out of the 18 samples, 15 were positive for beta-2 transferrin adn the leaks were validated surgically in 10 patients. Give leaks healed spontaneously with conservative management. Glucostix tests produced three false positive results from blood and nasal mucus contaminated fluid. Glucostix failed to detect another three CSF leaks resulting from false negative tests because of low CSF glucose levels. The Glucostix glucose test was nonspecific and insensitive compared with the beta-2 transferrin assay. CT failed to detect three of the 15 beta-2 transferrin-positive leaks but there were no false positive results. CT produced six negative results, of which three were false negatives. CONCLUSIONS: Glucose detection using Glucostix test strips is not recommended as a confirmatory test due to its lack of specificity and sensitivity. In the presence of a skull bas fracture on CT and a clinical CSF leak, there is no need for a further confirmatory test. In cases where a confirmatory test is needed, the beta-2 transferrin assay is the test of choice because of its high sensitivity and specificity.

Cerebrospinal Fluid Otorrhea↗

PubMed1994

Noninvasive method for the detection of bowel ischemia: computed tomography detection of absorbed iodinated contrast material.

RATIONALE AND OBJECTIVES: Although systemic absorption of enterically administered iohexol and its excretion in urine has been previously documented in rats with ischemic bowel, a practical and sensitive method of detecting urinary iohexol has not been available. We proposed to detect the presence of iohexol in the urine of rats with normal and ischemic bowel by use of a computed tomography (CT) number increase in the bladder with the use of CT. METHODS: Anesthetized rats (250 g) underwent either sham laparotomy (n = 6), ligation of two vascular arcades to the proximal jejunum (n = 5), ligation of six vascular arcades to the proximal jejunum (n = 6), or ligation of the superior mesenteric artery (n = 6). Rats were hydrated with saline (3.2 ml/hr intravenously). Each received a 3-ml enteric bolus of isotonic iohexol. Serial CT scans and plain film radiographs of the bladder were performed at 2, 4, and 6 hr to detect systemic absorption of contrast from the gut. Urine iohexol concentrations were measured by capillary electrophoresis. CT number and iohexol concentration were compared with evidence from plain film radiographs of bladder opacification. Intestinal ischemia was graded histologically. RESULTS: Histologic evidence of ischemia was present in all six-arcade and five of six superior mesenteric artery (SMA)-ligated animals. No animals in the control or two-arcade group showed evidence of bowel ischemia. Statistically significant increases (P < 0.05) in bladder density were demonstrated in the six-arcade and SMA-ligated groups. No statistical difference was noted between the two-arcade ligation and control groups. CONCLUSIONS: Experimental intestinal ischemia was reliably detected by bladder opacification after administration of enteric contrast. CT detection of systemic absorption of enteric iohexol was more sensitive than plain film radiographs and may be useful in the diagnosis of intestinal ischemia, although it may not be specific for ischemia.

Animals↗

PubMed1998

Diagnosis of schistosomiasis: antibody detection, with notes on parasitological and antigen detection methods.

Schistosomiasis remains a serious world-wide public health problem with a still unfulfilled need for routine cost-effective methods of diagnosis. Such methods are required not only for people in endemic areas, but increasingly for tourists who may have become infected during visits to such places. This article reviews the wide range of immunoassays and antigenic preparations that have been shown to have potential for diagnosis of schistosomiasis by the indirect method of antibody detection. Antigens in native form derived from cercariae, adult worms and eggs are considered, as well as schistosome antigens produced by recombinant DNA technology and the schistosome cross-reactive antigen, keyhole limpet haemocyanin (KLH). Respective advantages and disadvantages of antibody detection, circulating antigen detection and parasitological methods of diagnosis are analysed. It is suggested that due to the relative insensitivity of both parasitology and antigen detection, antibody detection methods could find increasing use in situations of low infection intensity.

Animals↗

PubMed2000

Detection of nanocolloids with flow-field flow fractionation and laser-induced breakdown detection.

A new nanosize colloid detection method comprised of flow-field flow fractionation (FFFF) with laser-induced breakdown detection (LIBD) is presented, which is capable of characterizing the colloid size distribution as well as determining the number density of each size fraction in very low concentrations. The method facilitates the detection of aquatic colloids particularly in the lower range of nanometer size (< 50 nm) with the sensitivity much higher than a laser light-scattering method (LLS), i.e., the lower ppb range. The method is tested with a mixture of polystyrene colloids in three different nominal sizes, 19, 50, and 102 nm, and the results are compared with those of the LLS method. For colloids of 19-nm diameter, the present method demonstrates the detection sensitivity over 3 orders of magnitude better than that of the LLS method. The limitation of the detection sensitivity arises from the surface bleeding of a ceramic frit overlying the separation channel of the used FFFF instrument.

Chemical Fractionation↗

PubMed2002

Genomic deletions of MSH2 and MLH1 in colorectal cancer families detected by a novel mutation detection approach.

Hereditary non-polyposis colorectal cancer is an autosomal dominant condition due to germline mutations in DNA-mismatch-repair genes, in particular MLH1, MSH2 and MSH6. Here we describe the application of a novel technique for the detection of genomic deletions in MLH1 and MSH2. This method, called multiplex ligation-dependent probe amplification, is a quantitative multiplex PCR approach to determine the relative copy number of each MLH1 and MSH2 exon. Mutation screening of genes was performed in 126 colorectal cancer families selected on the basis of clinical criteria and in addition, for a subset of families, the presence of microsatellite instability (MSI-high) in tumours. Thirty-eight germline mutations were detected in 37 (29.4%) of these kindreds, 31 of which have a predicted pathogenic effect. Among families with MSI-high tumours 65.7% harboured germline gene defects. Genomic deletions accounted for 54.8% of the pathogenic mutations. A complete deletion of the MLH1 gene was detected in two families. The multiplex ligation-dependent probe amplification approach is a rapid method for the detection of genomic deletions in MLH1 and MSH2. In addition, it reveals alterations that might escape detection using conventional diagnostic techniques. Multiplex ligation-dependent probe amplification might be considered as an early step in the molecular diagnosis of hereditary non-polyposis colorectal cancer.

Adaptor Proteins, Signal Transducing↗

PubMed1999

Hybrid Capture II-based human papillomavirus detection, a sensitive test to detect in routine high-grade cervical lesions: a preliminary study on 1518 women.

Hybrid Capture II (HC-II) is a commercial human papillomavirus (HPV) detection test designed to detect 18 HPV types divided into high-risk and low-risk groups. We have tested 1647 scrapes from 1518 unselected women attending routine cytological screening by this assay for the detection of histologically proven high-grade lesions. The reliability of this test was also evaluated on 117 fresh cone biopsy samples. HPV DNA has been detected in 400 scrapes (24.3%), 296 containing a high-risk HPV (18.0%). All the smears evocative of high-grade lesions were positive for high-risk HPV, and high-risk HPV were detected in all the 34 cases presenting a histologically proven high-grade lesion and in 68 (97.1%) of the 70 cone biopsy samples showing a high-grade lesion or an invasive carcinoma. Thus, the sensitivity was superior to the sensitivity of cytology (85.3%). Nevertheless, the quantitative approach provided by the HC-II assay for the assessment of the viral load could not clearly distinguish among cases with or without high-grade lesions. Thus this assay is recommended for the screening of high-grade lesions on a large scale, in association with classic cytology.

Adolescent↗

PubMed2004

Direct detection of renal function markers using microchip CE with pulsed electrochemical detection.

Creatinine, creatine, and uric acid are three important compounds that are measured in a variety of clinical assays, most notably for renal function. Traditional clinical assays for these compounds have focused on the use of enzymes or chemical reactions. Electrophoretic microchips have the potential to integrate separation power of capillary electrophoresis with devices that are small, portable, and have the speed of conventional sensors. The development of a microchip CE system for the direct detection of creatinine, creatine, and uric acid is presented. The device uses pulsed amperometric detection (PAD) to detect the nitrogen-containing compounds as well as the easily oxidizable uric acid. Baseline separation of creatinine, creatine and uric acid was achieved using 30 mM borate buffer (pH = 9.4) in less than 200 s. Linear calibration curves were obtained with limits of detection of 80 microM, 250 microM and 270 microM for creatinine, creatine and uric acid respectively. An optimization of the separation conditions and a comparison of PAD with other amperometric detection modes is also shown. Finally, analysis of a real urine sample is presented with validation of creatinine concentrations using a clinical assay kit based on the Jaffé reaction.

Biomarkers↗

PubMed2001

Early detection of hepatitis B surface antigen and detection of HBsAg mutants: a comparison of five assays.

BACKGROUND AND OBJECTIVES: Assays for the detection of hepatitis B surface antigen (HBsAg) face two challenges: the emergence of sensitive techniques for detection of hepatitis B virus DNA and the existence of HBsAg mutant hepatitis B viruses. We studied the sensitivity of five modern assays for detection of HBsAg. MATERIALS AND METHODS: The sensitivity of two mini robot-based assays--IMx HBsAg V2 and Vidas HBsAg--and three enzyme-linked immunosorbent assays (ELISAs) with microtitre plate format--Hepanostika Uni-Form II v1.2, Monolisa Ag HBs Plus and HBsAg Test System 3--was compared testing 11 HBsAg seroconversion series, serial dilutions prepared from three HBsAg standards and nine HBsAg mutant samples. RESULTS AND CONCLUSION: Overall, the IMx HBsAg V2 assay showed the highest sensitivity. It outperformed the Vidas HBsAg in analytical sensitivity and in detection of HBsAg mutants. Among the microtitre plate ELISAs, the Monolisa Ag HBs Plus outperformed the HBsAg Test System 3 in detection of HBsAg mutants and it surpassed the Hepanostika Uni-Form II v1.2 in analytical sensitivity.

Diagnostic Equipment↗