scholarly journals Automated Design of Probes for rRNA-Targeted FluorescenceIn SituHybridization Reveals the Advantages of Using Dual Probes for Accurate Identification

2014 ◽  
Vol 80 (16) ◽  
pp. 5124-5133 ◽  
Author(s):  
Erik S. Wright ◽  
L. Safak Yilmaz ◽  
Andrew M. Corcoran ◽  
Hatice E. Ökten ◽  
Daniel R. Noguera
Keyword(s):  
High Sensitivity ◽  
High Specificity ◽  
Target Group ◽  
Accurate Identification ◽  
Systematic Analysis ◽  
Single Probe ◽  
Common Technique ◽  
Generation Sequencing ◽  
Rrna Sequences

ABSTRACTFluorescencein situhybridization (FISH) is a common technique for identifying cells in their natural environment and is often used to complement next-generation sequencing approaches as an integral part of the full-cycle rRNA approach. A major challenge in FISH is the design of oligonucleotide probes with high sensitivity and specificity to their target group. The rapidly expanding number of rRNA sequences has increased awareness of the number of potential nontargets for every FISH probe, making the design of new FISH probes challenging using traditional methods. In this study, we conducted a systematic analysis of published probes that revealed that many have insufficient coverage or specificity for their intended target group. Therefore, we developed an improved thermodynamic model of FISH that can be applied at any taxonomic level, used the model to systematically design probes for all recognized genera of bacteria and archaea, and identified potential cross-hybridizations for the selected probes. This analysis resulted in high-specificity probes for 35.6% of the genera when a single probe was used in the absence of competitor probes and for 60.9% when up to two competitor probes were used. Requiring the hybridization of two independent probes for positive identification further increased specificity. In this case, we could design highly specific probe sets for up to 68.5% of the genera without the use of competitor probes and 87.7% when up to two competitor probes were used. The probes designed in this study, as well as tools for designing new probes, are available online (http://DECIPHER.cee.wisc.edu).

scholarly journals High-Sensitivity Radiocarbon Dating in the 50,000 to 70,000 bp Range without Isotopic Enrichment

Radiocarbon ◽  
1992 ◽  
Vol 34 (3) ◽  
pp. 351-359 ◽  
Author(s):  
Austin Long ◽  
Robert M. Kalin
Keyword(s):  
Memory Effect ◽  
Liquid Scintillation ◽  
High Sensitivity ◽  
Isotopic Enrichment ◽  
Counting System ◽  
Systematic Analysis ◽  
Critical Elements ◽  
Background Material ◽  
Acetylene Production

Calculations show the possibility of detecting 14C remaining after 10 or 11 decay half-lives in natural materials, such as wood, using commercially available liquid scintillation (LS) detectors. Assuming in-situ14C production has contributed insignificantly to the measured 14C, one can calculate finite ages approaching 70 ka. In practice, defensible finite age determinations involve careful considerations of several critical elements in the procedure. These critical elements are: 1) the integrity of the sample itself, in terms of younger contaminants and of in situ-produced 14C; 2) the availability of “dead” background material; 3) chemical blank in laboratory preparation of, in this case, benzene; and 4) stability of background and efficiency of the LS counting system. High-sensitivity 14C dating reveals a low level of memory effect 14C in benzene synthesized in the laboratory from anthracite or marble. This level of 14C activity is equivalent to that found in 53 ka old wood, and thus, is not distinguishable from petrochemical benzene used in routine dating. If careful control of laboratory conditions can maintain this 14C blank constant, reproducible dating beyond 53 ka would be possible. Although we have not completed a systematic analysis of the origins of memory effect, lithium reactors used in acetylene production and organic solvents in wood pretreatment are likely sources.

scholarly journals Correlation of Immunohistochemistry and Fluorescence in Situ Hybridization for HER-2 Assessment in Breast Cancer Patients: Single Centre Experience

2018 ◽  
Vol 6 (4) ◽  
pp. 593-599 ◽  
Author(s):  
Magdalena Bogdanovska-Todorovska ◽  
Slavica Kostadinova-Kunovska ◽  
Rubens Jovanovik ◽  
Blagica Krsteska ◽  
Goran Kondov ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cancer Patients ◽  
In Situ Hybridisation ◽  
False Negative ◽  
High Sensitivity ◽  
High Specificity ◽  
Fluorescence In Situ Hybridisation ◽  
Breast Cancer Patients ◽  
Her 2

BACKGROUND: Accurate assessment of HER-2 is imperative in selecting patients for targeted therapy. Most commonly used test methods for HER-2 are immunohistochemistry (IHC) and fluorescence in situ hybridisation (FISH). We evaluated the concordance between FISH and IHC for HER-2 in breast cancer samples using Food and Drug Administration approved tests.MATERIAL AND METHODS: Archived paraffin tissue blocks from 73 breast cancer patients were used. HER-2 immunostaining was performed using Ventana anti–HER-2 monoclonal antibody. The FISH assay was performed using PathVysion™ HER-2 DNA Probe Kit.RESULTS: Of the 73 cases 68.5% were IHC 0/1+, 15.07% were IHC 2+ and 16.44% were IHC 3+. Successful hybridisation was achieved in 72 cases. HER-2 FISH amplification was determined in 16.67% cases. Ten IHC 3+ and two IHC 2+ cases were FISH positive. Two of the IHC 3+ cases were FISH negative. Concordance rate was 100%, 18.18% and 83.33% for IHC 0/1+, 2+ and 3+ group, respectively. Total concordance was 84.72%, kappa 0.598 (p < 0.0001). The sensitivity of IHC in detecting IHC 2+ and IHC 3+ cases was 16.7% and 83.3%, and the specificity was 85% and 96.67%, respectively.CONCLUSION: The consistency between the methods was highest for IHC negative and lowest for IHC equivocal cases. The immunohistochemistry showed high sensitivity for IHC 2+/3+ cases and high specificity for IHC 3+ cases. Our results support the view that false-positive rather than false-negative IHC results are a problem with HER-2/IHC testing, and that IHC should be used as an initial screening test, but IHC 2+/ 3+ results should be confirmed by FISH.

Epstein-Barr Virus (EBV)-Encoded RNA: Automated In-Situ Hybridization (ISH) Compared with Manual ISH and Immunohistochemistry for Detection of EBV in Pediatric Lymphoproliferative Disorders

2009 ◽  
Vol 12 (3) ◽  
pp. 195-199 ◽  
Author(s):  
Naim K. Fanaian ◽  
Cynthia Cohen ◽  
Sandra Waldrop ◽  
Jennifer Wang ◽  
Bahig M. Shehata
Keyword(s):  
In Situ Hybridization ◽  
Predictive Value ◽  
Epstein Barr Virus ◽  
High Sensitivity ◽  
High Specificity ◽  
Lymphoproliferative Disorders ◽  
Barr Virus ◽  
Epstein Barr ◽  
Low Sensitivity

Detection of Epstein-Barr virus (EBV) may be achieved by various methods, including EBV-encoded RNA (EBER) in-situ hybridization (ISH) and immunohistochemistry (IHC) for latent membrane protein (LMP-1). We compared novel automated ISH and IHC techniques in pediatric lymphoproliferative disorders with results obtained by manual ISH. Thirty-seven pediatric cases previously studied by manual EBER ISH (including 18 EBER-positive, 15 EBER-negative, and 4 EBER-equivocal cases) were used for the study. Automated EBER ISH and automated LMP-1 IHC were performed using the BondMax autostainer and prediluted EBER probe and EBV cell surface 1 to 4 at 1:50 dilution, respectively. Results of each of the automated techniques for EBV detection were compared with results by manual EBER ISH. Compared with manual EBER ISH as the gold standard, automated ISH had a sensitivity and specificity of 94% and 69%, respectively, accuracy of 83%, positive predictive value (PPV) of 79%, and negative predictive value (NPV) of 90%. Automated IHC had a sensitivity of 44%, specificity of 93%, accuracy of 67%, PPV of 88%, and NPV of 59%. Automated ISH and IHC correlated significantly ( P < 0.045). Automated ISH is useful for diagnosis of EBV-related pediatric neoplasms, being easy to perform and interpret and requiring only the technologist's time to set up and having a high sensitivity and NPV. The automated IHC protocol is of too low sensitivity for routine use, although results show high specificity and PPV.

scholarly journals Title: LAMP Assays for the Simple and Rapid Detection of Clinically Important Urinary Pathogens Including the Detection of Resistance to 3rd Generation Cephalosporins

2021 ◽  
Author(s):  
Lalainasoa Odile RIVOARILALA ◽  
Victor JEANNODA ◽  
Tania CRUCITTI ◽  
Jean Marc COLLARD
Keyword(s):  
Simple Procedure ◽  
High Sensitivity ◽  
High Specificity ◽  
Urine Samples ◽  
Accurate Identification ◽  
Specificity And Sensitivity ◽  
Genes Encoding ◽  
Thermal Cycler ◽  
Tract Infections ◽  
Group 1

Abstract Background: Timely and accurate identification of uropathogens and determination of their antimicrobial susceptibility is paramount to the management of urinary tract infections (UTIs). The main objective of this study was to develop an assay using LAMP (Loop mediated isothermal amplification) technology for simple, rapid and sensitive detection of the most common bacteria responsible for UTIs, as well as for the detection of the most prevalent genes (encoding cefotaximases from CTX-M group 1) responsible for resistance to 3rd generation of cephalosporins. Method: We designed primers targeting Proteus mirabilis, while those targeting Escherichia coli, Klebsiella pneumoniae and Enterococcus faecalis and the CTX-M group 1 resistance gene were benchmarked from previous studies. The amplification reaction was carried out in a warm water bath for 60 min at 63±0.5 °C. The amplicons were revealed by staining with Sybr Green I. Specificity and sensitivity were determined using reference DNA extracts spiked in sterile urine samples. The analytical performance of the assays was evaluated directly on pellets of urine samples from patients suspected of UTI and compared with culture.Results: We found a high specificity (100%) for LAMP assays targeting the selected bacteria (P. mirabilis, E. coli, K. pneumoniae, E. faecalis) and the CTX-M group 1 when using DNA extracts spiked in urine samples. The sensitivities of the assays were around 1.5 103 Colony Forming Units (CFU) /mL corresponding to the cut-off value used to define bacteriuria or UTIs in patients with symptoms. Out of 161 urine samples tested, using culture as gold standard, we found a sensitivity of the LAMP techniques ranging from 96 to 100 % and specificity from 95 to 100 %.Conclusion: We showed that the LAMP assays were simple and fast. The tests showed high sensitivity and specificity using a simple procedure for DNA extraction. In addition, the assays could be performed without the need of an expensive device such as a thermal cycler. These LAMP assays could be useful as an alternative or a complementary tool to culture reducing the time to diagnosis and guiding for more effective treatment of UTIs but also as a powerful diagnostic tool in resource-limited countries where culture is not available in primary health care structures.

scholarly journals Market Perspectives and Future Fields of Application of Odor Detection Biosensors—A Systematic Analysis

Proceedings ◽  
2020 ◽  
Vol 60 (1) ◽  
pp. 40
Author(s):  
Johannes Full ◽  
Lukas Delbrück ◽  
Alexander Sauer ◽  
Robert Miehe
Keyword(s):  
Health And Safety ◽  
High Sensitivity ◽  
High Specificity ◽  
Systematic Analysis ◽  
Odor Detection ◽  
Application Potential ◽  
Development Goals ◽  
Performance Profiles ◽  
And Performance ◽  
Application Fields

The technological advantages that biosensors have over conventional technical sensors for odor detection have not yet been comprehensively analyzed. However, this is necessary for assessing their suitability for specific fields of application as well as their improvement and development goals. In this paper specific market potentials of biosensors for odor detection are identified by applying a tailored methodology that enables the derivation and systematic comparison of both the performance profiles of biosensors as well as the requirement profiles for various application fields. Therefore, the fulfillment of defined requirements is evaluated for biosensors by means of 16 selected technical criteria in order to determine a specific performance profile. Further, a selection of application fields for odor detection sensors is derived to compare the importance of the criteria for each of the fields, leading to market-specific requirement profiles. The analysis reveals that the requirement criteria considered to be the most important ones across all application fields are high specificity, high selectivity, high repeat accuracy, high resolution, high accuracy, and high sensitivity. All these criteria, except for the repeat accuracy, can potentially be better met by biosensors than by technical sensors, according to the results obtained. Therefore, biosensor technology in general has a high application potential for all the areas of application under consideration. Health and safety applications especially are considered to have high potential for biosensors due to their correspondence between requirement and performance profiles.

scholarly journals Artificial Intelligence Can Cut Costs While Maintaining Accuracy in Colorectal Cancer Genotyping

2021 ◽  
Vol 11 ◽  
Author(s):  
Alec J. Kacew ◽  
Garth W. Strohbehn ◽  
Loren Saulsberry ◽  
Neda Laiteerapong ◽  
Nicole A. Cipriani ◽  
...  
Keyword(s):  
Artificial Intelligence ◽  
Colorectal Cancer ◽  
Next Generation Sequencing ◽  
Diagnostic Accuracy ◽  
Metastatic Colorectal Cancer ◽  
High Sensitivity ◽  
High Specificity ◽  
Next Generation ◽  
First Line ◽  
Generation Sequencing

Rising cancer care costs impose financial burdens on health systems. Applying artificial intelligence to diagnostic algorithms may reduce testing costs and avoid wasteful therapy-related expenditures. To evaluate the financial and clinical impact of incorporating artificial intelligence-based determination of mismatch repair/microsatellite instability status into the first-line metastatic colorectal carcinoma setting, we developed a deterministic model to compare eight testing strategies: A) next-generation sequencing alone, B) high-sensitivity polymerase chain reaction or immunohistochemistry panel alone, C) high-specificity panel alone, D) high-specificity artificial intelligence alone, E) high-sensitivity artificial intelligence followed by next generation sequencing, F) high-specificity artificial intelligence followed by next-generation sequencing, G) high-sensitivity artificial intelligence and high-sensitivity panel, and H) high-sensitivity artificial intelligence and high-specificity panel. We used a hypothetical, nationally representative, population-based sample of individuals receiving first-line treatment for de novo metastatic colorectal cancer (N = 32,549) in the United States. Model inputs were derived from secondary research (peer-reviewed literature and Medicare data). We estimated the population-level diagnostic costs and clinical implications for each testing strategy. The testing strategy that resulted in the greatest project cost savings (including testing and first-line drug cost) compared to next-generation sequencing alone in newly-diagnosed metastatic colorectal cancer was using high-sensitivity artificial intelligence followed by confirmatory high-specificity polymerase chain reaction or immunohistochemistry panel for patients testing negative by artificial intelligence ($400 million, 12.9%). The high-specificity artificial intelligence-only strategy resulted in the most favorable clinical impact, with 97% diagnostic accuracy in guiding genotype-directed treatment and average time to treatment initiation of less than one day. Artificial intelligence has the potential to reduce both time to treatment initiation and costs in the metastatic colorectal cancer setting without meaningfully sacrificing diagnostic accuracy. We expect the artificial intelligence value proposition to improve in coming years, with increasing diagnostic accuracy and decreasing costs of processing power. To extract maximal value from the technology, health systems should evaluate integrating diagnostic histopathologic artificial intelligence into institutional protocols, perhaps in place of other genotyping methodologies.

scholarly journals Biomarkers for detection and surveillance of bladder cancer

2013 ◽  
Vol 2 (3) ◽  
pp. 212 ◽  
Author(s):  
Lorne I. Budman ◽  
Wassim Kassouf ◽  
Jordan R. Steinberg
Keyword(s):  
Bladder Cancer ◽  
Bladder Tumour ◽  
Transitional Cell ◽  
High Sensitivity ◽  
High Specificity ◽  
Urinary Cytology ◽  
Term Survival ◽  
Common Cancer ◽  
Health Canada

Introduction: Bladder cancer is the fourth most common cancer in men and the ninth most common cancer in women in Canada. Early detection of tumours is essential for improved prognosis and long-term survival. The standard method for detection and surveillance is cystoscopy together with urine cytology. Cystoscopy is relatively sensitive but is expensive and invasive. Urinary cytology is a noninvasive method that has poor sensitivity but high specificity; it is relied on for the detection of carcinoma in situ. Currently, several urinary based bladder tumour biomarkers with USFDA/Health Canada approval are available commercially, but none have been widely adopted by urologists despite their offering high sensitivity and/or specificity. We present here a review of recent studies evaluating 7 commercial biomarker assays for the detection and/or surveillance of bladder cancer.Results: Sensitivity and specificity ranges, respectively, for each marker were reported as follows: BTA Stat (Polymedco), 52.5%–78.0% and 69.0%–87.1%; BTA Trak (Polymedco), 51%–100% and 73%–92.5%; cytology, 12.1%–84.6% and 78.0%–100%; hematuria dipstick, 47.0%–92.6% and 51.0%–84.0%; NMP22 Bladder Cancer Test (Matritech), 34.6%–100% and 60.0%–95.0%; NMP22 BladderChek (Matritech), 49.5%–65.0% and 40.0%–89.8%;ImmunoCyt/uCyt+ (DiagnoCure), 63.3%–84.9% and 62.0%–78.1%; ImmunoCyt/uCyt+ and cytology, 81.0%–89.3% and 61.0%–77.7%; and UroVysion (Abbott Molecular)/florescence in situ hybridization, 68.6%–100% and 65.0%–96.0%.Conclusion: We find that no currently available bladder cancer urinary marker is sensitive enough to eliminate the need for cystoscopy. In addition, cytology remains integral to the detection of occult cancer. However, owing to their relatively high sensitivities, these markers may be used to extend the period between cystoscopies in the surveillance of patients with transitional cell carcinoma. Further study is required to determine which markers, alone or in panel, would best accomplish this.

scholarly journals Detection of aerobe–anaerobe mixed infection by metagenomic next-generation sequencing in an adult suffering from descending necrotizing mediastinitis

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Jing Duan ◽  
Chuncheng Zhang ◽  
Xiaoshuang Che ◽  
Juanjuan Fu ◽  
Feng Pang ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Mixed Infection ◽  
Clinical Laboratory ◽  
High Sensitivity ◽  
Future Trend ◽  
Descending Necrotizing Mediastinitis ◽  
Next Generation ◽  
Accurate Identification ◽  
Drainage Fluid ◽  
Generation Sequencing

Abstract Background Descending necrotizing mediastinitis (DNM) is one of the most virulent forms of mediastinitis. The main causes of high mortality in DNM are believed to stem from difficulty and delay in the diagnosis. Fast and accurate identification of pathogens is important for the treatment of these patients. Metagenomics next-generation sequencing (mNGS) is a powerful tool to identify all kinds of pathogens, especially for rare and complex infections. Case presentation A 64-year-old male patient was admitted to the intensive care unit (ICU) with unconsciousness, dyspnea, and swelling in the mandible and neck. Computed tomography (CT) scan results combined with clinical laboratory examination indicated DNM. Vancomycin and imipenem were used, and vacuum sealing drainage was applied for debridement and drainage of the infected area. The positive mNGS results of drainage fluid confirmed the presence of mixed infection caused by Streptococcus anginosus, Prevotella oris, and several other anaerobes. The antibiotics were adjusted to piperacillin/tazobactam and tinidazole according to the mNGS results and antimicrobial susceptibility testing of cultured pathogens. After 11 days of antibiotic therapy, the infection symptoms of the neck and mediastinum improved, and the patient was transferred out of the ICU on the 26th day after negative result of drainage fluid culture. Conclusion This case suggested that mNGS is a promising technology for precise and fast pathogens identification with high sensitivity, which may guide the diagnosis of infectious diseases in the future trend.

scholarly journals LAMP assays for the simple and rapid detection of clinically important urinary pathogens including the detection of resistance to 3rd generation cephalosporins

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Lalainasoa Odile Rivoarilala ◽  
Jeannoda Victor ◽  
Tania Crucitti ◽  
Jean Marc Collard
Keyword(s):  
Simple Procedure ◽  
High Sensitivity ◽  
High Specificity ◽  
Urine Samples ◽  
Accurate Identification ◽  
Specificity And Sensitivity ◽  
Genes Encoding ◽  
Thermal Cycler ◽  
Tract Infections ◽  
Group 1

Abstract Background Timely and accurate identification of uropathogens and determination of their antimicrobial susceptibility is paramount to the management of urinary tract infections (UTIs). The main objective of this study was to develop an assay using LAMP (Loop mediated isothermal amplification) technology for simple, rapid and sensitive detection of the most common bacteria responsible for UTIs, as well as for the detection of the most prevalent genes (encoding cefotaximases from CTX-M group 1) responsible for resistance to 3rd generation of cephalosporins. Method We designed primers targeting Proteus mirabilis, while those targeting Escherichia coli, Klebsiella pneumoniae and Enterococcus faecalis and the CTX-M group 1 resistance gene were benchmarked from previous studies. The amplification reaction was carried out in a warm water bath for 60 min at 63 ± 0.5 °C. The amplicons were revealed by staining with Sybr Green I. Specificity and sensitivity were determined using reference DNA extracts spiked in sterile urine samples. The analytical performance of the assays was evaluated directly on pellets of urine samples from patients suspected of UTI and compared with culture. Results We found a high specificity (100%) for LAMP assays targeting the selected bacteria (P. mirabilis, E. coli, K. pneumoniae, E. faecalis) and the CTX-M group 1 when using DNA extracts spiked in urine samples. The sensitivities of the assays were around 1.5 103 Colony Forming Units (CFU) /mL corresponding to the cut-off value used to define bacteriuria or UTIs in patients with symptoms. Out of 161 urine samples tested, using culture as gold standard, we found a sensitivity of the LAMP techniques ranging from 96 to 100% and specificity from 95 to 100%. Conclusion We showed that the LAMP assays were simple and fast. The tests showed high sensitivity and specificity using a simple procedure for DNA extraction. In addition, the assays could be performed without the need of an expensive device such as a thermal cycler. These LAMP assays could be useful as an alternative or a complementary tool to culture reducing the time to diagnosis and guiding for more effective treatment of UTIs but also as a powerful diagnostic tool in resource-limited countries where culture is not available in primary health care structures.

scholarly journals Market Perspectives and Future Fields of Application of Odor Detection Biosensors within the Biological Transformation—A Systematic Analysis

Biosensors ◽  
2021 ◽  
Vol 11 (3) ◽  
pp. 93
Author(s):  
Johannes Full ◽  
Yannick Baumgarten ◽  
Lukas Delbrück ◽  
Alexander Sauer ◽  
Robert Miehe
Keyword(s):  
Health And Safety ◽  
High Sensitivity ◽  
High Specificity ◽  
Systematic Analysis ◽  
Odor Detection ◽  
Biological Transformation ◽  
Performance Profiles ◽  
And Performance ◽  
Application Fields ◽  
Safety Applications

The technological advantages that biosensors have over conventional technical sensors for odor detection and the role they play in the biological transformation have not yet been comprehensively analyzed. However, this is necessary for assessing their suitability for specific fields of application as well as their improvement and development goals. An overview of biological basics of olfactory systems is given and different odor sensor technologies are described and classified in this paper. Specific market potentials of biosensors for odor detection are identified by applying a tailored methodology that enables the derivation and systematic comparison of both the performance profiles of biosensors as well as the requirement profiles for various application fields. Therefore, the fulfillment of defined requirements is evaluated for biosensors by means of 16 selected technical criteria in order to determine a specific performance profile. Further, a selection of application fields, namely healthcare, food industry, agriculture, cosmetics, safety applications, environmental monitoring for odor detection sensors is derived to compare the importance of the criteria for each of the fields, leading to market-specific requirement profiles. The analysis reveals that the requirement criteria considered to be the most important ones across all application fields are high specificity, high selectivity, high repeat accuracy, high resolution, high accuracy, and high sensitivity. All these criteria, except for the repeat accuracy, can potentially be better met by biosensors than by technical sensors, according to the results obtained. Therefore, biosensor technology in general has a high application potential for all the areas of application under consideration. Health and safety applications especially are considered to have high potential for biosensors due to their correspondence between requirement and performance profiles. Special attention is paid to new areas of application that require multi-sensing capability. Application scenarios for multi-sensing biosensors are therefore derived. Moreover, the role of biosensors within the biological transformation is discussed.

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