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PubMedDate not supplied

A pilot study for a screening trial of cervical fluorescence spectroscopy.

Fluorescence spectroscopy is a promising technology for detection of epithelial precancers and cancers. In preparation for a multicenter phase II screening trial, a pilot trial was conducted to test data collection and patient examination procedures, use data forms, time procedures, and identify problems with preliminary data analysis. Women 18 years of age and older underwent a questionnaire, a complete history, and a physical examination, including a pan-colposcopy of the lower genital tract. A fiber-optic probe measured fluorescence excitation-emission matrices at 1-3 cervical sites for 58 women. The data collection procedures, data forms, and procedure times worked well, although collection times for all the clinical data take an average of 28 min. The clinical team followed procedures well, and the data could be retrieved from the database at all sites. The multivariate analysis algorithm correctly identified squamous normal tissue 99% of the time and columnar normal tissue only 7%. The assessment of ploidy from monolayer samples was not accurate in this small sample. The study was successful as a pilot trial. We learned who participated, who withdrew, how often abnormalities were present, and that algorithms that have worked extremely well in previous studies do not work as well when a few study parameters are changed. The current algorithm for diagnosis identified squamous normal tissue very accurately and did less well for columnar normal tissue. Inflammation may be an explanation for this phenomenon. Fluorescence spectroscopy is a promising technology for the detection of epithelial precancers and cancers. The screening trial of fluorescence and reflectance spectroscopy was successful.

Adult↗

PubMed2002

Magnetic resonance imaging in detection and functional assessment of coronary artery disease.

The past few years have brought significant improvements in the field of cardiovascular magnetic resonance imaging (MRI), which evolved from an experimental technique to a clinically accepted method of coronary artery disease detection (stress MRI) and viability assessment. In this article, we describe current MRI technology for detection and functional assessment of ischemia, such as dobutamine/atropine MRI, perfusion techniques, viability, and flow reserve in native coronary arteries and grafts. With further refinement in the technology, wide acceptance of cardiovascular MRI is anticipated in clinical practice.

Atropine↗

PubMed2000

The ligase chain reaction as a primary screening tool for the detection of culture positive tuberculosis.

BACKGROUND: The ligase chain reaction Mycobacterium tuberculosis assay uses ligase chain reaction technology to detect tuberculous DNA sequences in clinical specimens. A study was undertaken to determine its sensitivity and specificity as a primary screening tool for the detection of culture positive tuberculosis. METHODS: The study was conducted on 2420 clinical specimens (sputum, bronchoalveolar lavage fluid, pleural fluid, urine) submitted for primary screening for Mycobacterium tuberculosis to a regional medical microbiology laboratory. Specimens were tested in parallel with smear, ligase chain reaction, and culture. RESULTS: Thirty nine patients had specimens testing positive by the ligase chain reaction assay. Thirty two patients had newly diagnosed tuberculosis, one had a tuberculosis relapse, three had tuberculosis (on antituberculous therapy when tested), and three had healed tuberculosis. In the newly diagnosed group specimens were smear positive in 21 cases (66%), ligase chain reaction positive in 30 cases (94%), and culture positive in 32 cases (100%). Using a positive culture to diagnose active tuberculosis, the ligase chain reaction assay had a sensitivity of 93.9%, a specificity of 99.8%, a positive predictive value of 83.8%, and a negative predictive value of 99.9%. CONCLUSIONS: This study is the largest clinical trial to date to report the efficacy of the ligase chain reaction as a primary screening tool to detect Mycobacterium tuberculosis infection. The authors conclude that ligase chain reaction is a useful primary screening test for tuberculosis, offering speed and discrimination in the early stages of diagnosis and complementing traditional smear and culture techniques.

Adolescent↗

PubMed2005

Shaking the tree: mapping complex disease genes with linkage disequilibrium.

Much effort and expense are being spent internationally to detect genetic polymorphisms contributing to susceptibility to complex human disease. Concomitantly, the technology for detecting and genotyping single nucleotide polymorphisms (SNPs) has undergone rapid development, yielding extensive catalogues of these polymorphisms across the genome. Population-based maps of the correlations amongst SNPs (linkage disequilibrium) are now being developed to accelerate the discovery of genes for complex human diseases. These genomic advances coincide with an increasing recognition of the importance of very large sample sizes for studying genetic effects. Together, these new genetic and epidemiological data hold renewed promise for the identification of susceptibility genes for complex traits. We review the state of knowledge about the structure of the human genome as related to SNPs and linkage disequilibrium, discuss the potential applications of this knowledge to mapping complex disease genes, and consider the issues facing whole genome association scanning using SNPs.

Chromosome Mapping↗

PubMed2001

Electrospray ionization mass spectrometry-based genotyping: an approach for identification of single nucleotide polymorphisms.

The high frequency of single nucleotide polymorphisms (SNPs) in the human genome makes them ideal genetic markers for mapping, diagnosing disease-related alleles, and identifying SNPs that contribute to drug response differences between individuals. Here we report a novel assay utilizing a single nucleotide primer extension (SNuPE) and electrospray ionization mass spectrometry (ESI-MS) detection for the analysis of SNPs. In contrast to most SNuPE genotyping technologies that detect the extended primer product, the novel Survivor assay detects the unreacted dideoxynucleotides (ddNTPs) remaining or surviving in solution following a SNuPE. This assay involves a simple analysis of the same four ddNTP analytes, regardless of the SNP being investigated, and either single or double-stranded DNA can be used to genotype a SNP, without any labeling requirements of the ddNTPs or oligonucleotide primers. We have tested and blindly validated the Survivor assay by genotyping the C/T SNP at -857 of the human TNFalpha promoter gene. The results obtained are in agreement with the control sequencing data. The results demonstrate that the homogeneous Survivor assay with ESI-MS detection offers advantages in simplicity, accuracy, specificity, and sensitivity. Additional advantages of the method include enhanced hybridization efficiencies in this solution-phase assay and the elimination of immobilized primers for the isolation of single-stranded DNA. With a one-well reaction and an automation platform being developed, the Survivor assay provides a powerful new tool for large-scale SNP analysis and screening.

Base Sequence↗

PubMed2005

An airport cargo inspection system based on X-ray and thermal neutron analysis (TNA).

A cargo inspection system incorporating a high-resolution X-ray imaging system with a material-specific detection system based on Ancore Corporation's patented thermal neutron analysis (TNA) technology can detect bulk quantities of explosives and drugs concealed in trucks or cargo containers. The TNA process utilises a 252Cf neutron source surrounded by a moderator. The neutron interactions with the inspected object result in strong and unique gamma-ray signals from nitrogen, which is a key ingredient in modern high explosives, and from chlorinated drugs. The TNA computer analyses the gamma-ray signals and automatically determines the presence of explosives or drugs. The radiation source terms and shielding design of the facility are described. For the X-ray generator, the primary beam, leakage radiation, and scattered primary and leakage radiation were considered. For the TNA, the primary neutrons and tunnel scattered neutrons as well as the neutron-capture gamma rays were considered.

Aircraft↗

PubMed2001

Using single nucleotide polymorphisms as a means to understanding the pathophysiology of asthma.

Asthma is the most common chronic childhood disease in the developed nations, and is a complex disease that has high social and economic costs. Studies of the genetic etiology of asthma offer a way of improving our understanding of its pathogenesis, with the goal of improving preventive strategies, diagnostic tools, and therapies. Considerable effort and expense have been expended in attempts to detect specific polymorphisms in genetic loci contributing to asthma susceptibility. Concomitantly, the technology for detecting single nucleotide polymorphisms (SNPs) has undergone rapid development, extensive catalogues of SNPs across the genome have been constructed, and SNPs have been increasingly used as a method of investigating the genetic etiology of complex human diseases. This paper reviews both current and potential future contributions of SNPs to our understanding of asthma pathophysiology.

Animals↗

PubMed1999

[Mycobacteria--methods to meet the CDC guidelines].

In response to the guidelines from the Centers for Disease Control and Prevention, Atlanta, U.S.A. (1994), a considerable effort has been focused on the development and application of rapid laboratory testings for Mycobacterium tuberculosis. Concerning the first goal, "acid-fast bacillus smear results should be reported within 24 hr of specimen collection", most laboratories latently have an ability to perform unless the specimens will be processed in a lump. However, the second goal, "isolation and identification of M. tuberculosis should be within 10 to 14 days", is almost impossible for most Japanese laboratories where the radiometric Bactec 460 system is not applicable. In recent years, several nonradiometric, automated or semiautomated mycobacterial culture systems are developed and applied. To date, the evaluated results indicate that the newly developed mycobacterial culture systems with Middlebrook 7H9 broth can report positive with means of 17 to 20 day incubation, 3 to 6 days behind the CDC guidelines. Finally, regarding the third goal, "susceptibility test results should be within a total of 15 to 30 days", our developed microdilution test method can report quantitative minimum inhibitory concentrations (MICs) for M. tuberculosis after 7 day incubation. Overall, our procedures are very close to the CDC guidelines, except for isolation and identification of M. tuberculosis. The remaining key to fulfil the guidelines is a rapid detection, that is, culture media which can support rapid mycobacterial growth and advanced technology to detect attenuated growth of M. tuberculosis.

Bacteriological Techniques↗

PubMed2006

Blinded, multicenter comparison of methods to detect a drug-resistant mutant of human immunodeficiency virus type 1 at low frequency.

We determined the abilities of 10 technologies to detect and quantify a common drug-resistant mutant of human immunodeficiency virus type 1 (lysine to asparagine at codon 103 of the reverse transcriptase) using a blinded test panel containing mutant-wild-type mixtures ranging from 0.01% to 100% mutant. Two technologies, allele-specific reverse transcriptase PCR and a Ty1HRT yeast system, could quantify the mutant down to 0.1 to 0.4%. These technologies should help define the impact of low-frequency drug-resistant mutants on response to antiretroviral therapy.

Anti-HIV Agents↗

PubMed1991

Detection of Escherichia coli rRNA using target amplification and time-resolved fluorescence detection.

The development of technology to increase the sensitivity and speed of detection of bacterial pathogens in samples is important for diagnosis and monitoring of illness. We have developed a sensitive and rapid method for the detection of bacteria, using Escherichia coli as a model, which combines transcription-based target amplification with a bead-based sandwich hybridization assay using rare earth metal chelate labelled probes and time-resolved fluorescence detection. Using these methods as little as 100 copies (0.00016 attomoles) of purified native Escherichia coli rRNA or just one bacterial cell in a spiked sample could be detected. These results demonstrate that amplification of rRNA by transcription-based amplification and detection by time-resolved fluorescence provide a sensitive technology for the direct detection of micro-organisms without the requirement for prior cultivation.

Base Sequence↗

PubMedDate not supplied

Proteins, recognition networks and developing interfaces for macromolecular biosensing.

Genomics and proteomics discovery is leading to the identification of all proteins and to the opportunity, and challenge, to reveal the protein recognition networks that drive virtually all biological processes. Over the past decade, biosensors have emerged as a key technology for detection and analysis of biomolecular interactions. An important limitation in developing such biosensors is that the focus has been mainly on sensor platforms, the transducing hardware that converts interaction signals into recorded data, without adequately considering the role of molecular interfaces, the elements of sensors that interact with analytes to produce signals. We have investigated this alternative focus by identifying and, where necessary, designing molecular interfaces that will more effectively drive new biosensor development and utilization in biomedical and biotechnological investigations. Here we describe our recent studies of coiled coil and lipid bilayer interfaces and the potential to use these to expand sensing technologies for multiplexed target detection and analysis in increasingly biologically relevant membrane like environments.

Amino Acid Motifs↗

PubMed2001

Characterization of the groESL operon in Listeria monocytogenes: utilization of two reporter systems (gfp and hly) for evaluating in vivo expression.

The ability of intracellular pathogens to sense and adapt to the hostile environment of the host is an important factor governing virulence. We have sequenced the operon encoding the major heat shock proteins GroES and GroEL in the gram-positive food-borne pathogen Listeria monocytogenes. The operon has a conserved orientation in the order groES groEL. Upstream of groES and in the opposite orientation is a gene encoding a homologue of the Bacillus subtilis protein YdiL, while downstream of groEL is a gene encoding a putative bile hydrolase. We used both reverse transcriptase-PCR (RT-PCR) and transcriptional fusions to the UV-optimized Aequorea victoria green fluorescent protein (GFP(UV)) to analyze expression of groESL under various environmental stress conditions, including heat shock, ethanol stress, and acid shock, and during infection of J774 mouse macrophage cells. Strains harboring GFP(UV) transcriptional fusions to the promoter region of groESL demonstrated a significant increase in fluorescence following heat shock that was detected by both fluorimetry and fluorescence microscopy. Using both RT-PCR and GFP technology we detected expression of groESL following internalization by J774 cells. Increased intracellular expression of dnaK was also determined using RT-PCR. We have recently described a system which utilizes L. monocytogenes hemolysin as an in vivo reporter of gene expression within the host cell phagosome (C. G. M. Gahan and C. Hill, Mol. Microbiol. 36:498-507, 2000). In this study a strain was constructed in which hemolysin expression was placed under the control of the groESL promoter. In this strain hemolysin expression during infection also confirms transcription from the groESL promoter during J774 and murine infection, albeit at lower levels than the known virulence factor plcA.

Animals↗

PubMed2005

New frontiers in endoscopic imaging.

Gastrointestinal (GI) tract malignancies have a tremendous impact on society. Colorectal cancer is the second leading cause of cancer death in the United States and accounts for 10% of all cancer deaths. Significant research efforts are being directed toward using the interaction of light and tissue to detect pre-cancerous lesions of the GI tract. This article reviews the current status of various experimental optical technologies to detect pre-cancerous changes in the GI tract and focuses on the clinical applications of these technologies for the practicing gastroenterologist.

Diagnostic Imaging↗

PubMed2002

[Analysis of gene expression at the entire genome scale--applications in biological sciences and medicine].

Microarrays are one of the latest breakthroughs in experimental molecular biology, which allow monitoring of gene expression for tens of thousands of genes in parallel and are already producing huge amounts of valuable data. Microarray RNA expression on a genome-wide range is now a proven technology, although the idea of analysis of expression many genes in one sample is not new. Since the development of clone printing technology and oligonucleotide synthesis allowed to produce high density microarray. In this publication we provide the information about the technology, available detection systems and data analysis software. Comprehensive review of current or fundamental papers using microarray technology application in immunology, rheumatoid arthritis, oncology, cystis fibrosis research, primary pulmonary hypertension, psychiatry, and allergic airways inflammation is also included.

Animals↗

PubMed2004

Detection of MYCN amplification and chromosome 1p36 loss in neuroblastoma by cDNA microarray comparative genomic hybridization.

BACKGROUND: In the last decade, microarray technology has been extensively used to evaluate gene expression profiles and genome imbalances. We have developed a microarray-based comparative genomic hybridization (CGH) approach to identify MYCN gene amplification and 1p36 chromosome loss, two markers of tumor aggressiveness in neuroblastoma. AIM: The aim was to use microarray CGH technology to detect the two major prognostic markers for neuroblastoma, MYCN amplification and 1p36 chromosome deletion, in neuroblastoma patients and, therefore, confirm the usefulness of this approach in this cancer. METHODS: DNA was purified from 16 tumors containing at least 90% malignant neuroblasts and collected at the onset of disease. Pooled fluorescent-labeled reference and neuroblastoma tumor genomic DNA was hybridized to epoxide-coated glass slides on laboratory-made complementary DNA microarray. The microarray contained cDNA mapped at the 1p36.33-36.1 chromosomal region and MYCN gene. cDNA from the 2q33-q34 and 12p13 chromosomes was used as a control and Arabidopsis thaliana DNA was spotted to control unspecific hybridization. Fluorescence in situ hybridization analysis was also performed to validate results from the microarray CGH. RESULTS: Both MYCN amplification and 1p36 chromosome deletion were detected by microarray CGH. The sensitivity and specificity for 1p36 loss detection were 66.7% and 90.0%, respectively. The method had a sensitivity of 66.7% and specificity of 90.9% to detect MYCN amplification. DISCUSSION: Our results demonstrated that the microarray CGH can be efficiently applied to study DNA gain and loss of specific chromosome regions.

Cell Nucleus↗

PubMed2006

A novel generic dipstick-based technology for rapid and precise detection of tetracycline, streptogramin and macrolide antibiotics in food samples.

Excessive use of antibiotics in veterinary medicine and as growth promoters in stock farming has been associated with the dramatically increasing prevalence of multidrug-resistant human pathogenic bacteria. European community legislators have therefore restricted the veterinary use of antibiotics and banned them as growth-promoting food additives in stock breeding (1831/2003/EC). The monitoring of such legislation requires technology for precise and straightforward on-site quantification of antibiotics in farm samples and food products without the need for extensive laboratory equipment and trained personnel. Capitalizing on bacterial transcriptional regulators (TetR, PIP, E), which are dose-dependently released from their cognate operators (tetO, PIR, ETR) upon binding of specific classes of antibiotics (tetracycline, streptogramins, macrolides) we have designed an easy-to-handle dipstick-based assay for detection of antibiotic levels in serum, meat and milk whose detection limits are up to 40-fold below licensed threshold values. The generic dipstick consists of either nitrocellulose, nylon or polyvinylidenfluorid (PVDF) membrane strips coated with streptavidin and immobilized biotinylated operator DNA, which acts as capture DNA to bind hexa-histidine (His(6))-tagged bacterial biosensors. Antibiotics present in specific samples triggered the dose-dependent release of the capture DNA-biosensor interaction, which, after dipping into two different solutions, results in a correlated conversion of a chromogenic substrate by a standard His(6)-targeted enzyme complex. This can be quantified by comparison of the dipstick to a standardized color scale or by assessing the terminal solution at 450nm. As demonstrated using serum, meat and milk samples spiked with 14 different antibiotics, the dipstick technology provided sensitive detection in a rapid assay format, and could be employed to monitor non-authorized use of antibiotics and to discover novel antibiotics.

Animals↗

PubMedDate not supplied

Circulating tumor cells and serum tumor biomarkers in small cell lung cancer.

Lung cancer accounts for approximately 30% of all cancer mortalities in the United States. Small cell lung cancer (SCLC), which is an aggressive malignancy with frequent and early metastases, accounts for about 15% of all of the lung cancer cases with a dismal 5-year survival rate of < 5% with current standard therapies. Early detection of SCLC is challenging, in part due to the lack of adequate serum tumor markers. The goal of this review is to summarize the current knowledge of circulating tumor cells and serum biomarkers in small cell lung cancer. The role of circulating tumor cells in prognostication is controversial, but may be better defined with advancing technologies of detection of such cells with higher precision, and improved clinico-pathological correlations. The current knowledge on the known serum cytokines and tumor biomarkers of SCLC, such as CEA, chromogranin-A and neuron-specific enolase will be presented. Serum cytokines, such as vascular endothelial growth factor (VEGF), stem cell factor (SCF) and hepatocyte growth factor/scatter factor (HGF/SF) are also discussed. New findings in the search for novel diagnostic and therapeutic molecular markers using the emerging genomics and proteomics technologies are emphasized. It is our hope that validation of these new research platforms and technologies will result in improved early detection, prognostication and finally treatment of SCLC with potential novel molecularly-targeted therapeutics.

Biomarkers, Tumor↗

PubMedDate not supplied

Early detection of secondary caries using quantitative, light-induced fluorescence.

The authors hypothesize that the arrestment and remineralization of these lesions could be improved if secondary caries could be detected and monitored at earlier stages. Traditional diagnostic techniques detect secondary caries when it is relatively advanced and when significant tissue has been lost. This in vitro study evaluated the Quantitative Light-induced Fluorescence (QLF) system for detecting and monitoring demineralization surrounding tooth-colored restorations. This investigation was divided into three studies. The first study evaluated QLF for the detection and measurement of chemically induced lesions surrounding resin composite restorations. The second study evaluated QLF for the detection of demineralization around different tooth-colored restorations (glass ionomer, resin composite, compomer and smart material) created in a microbial caries model. Finally, the third study tested whether QLF was effective at detecting early wall lesions adjacent to resin composite restorations. Data from Study 1 demonstrated the potential for QLF to detect very early secondary caries and to distinguish between the different stages of early demineralization. Study 2 confirmed the potential for QLF to detect early secondary caries created by cariogenic bacteria and concluded that the four types of materials differed in their ability to prevent secondary caries in this model, with the glass ionomer being the most effective and the non-fluoride releasing composite performing the worst, which allowed for the development of larger lesions. The results of Study 3 suggest a potential application of the QLF system to detect early wall lesions. Data from this investigation strongly suggest that QLF is a potentially viable technology to detect and monitor early secondary caries.

Analysis of Variance↗