scholarly journals Automation of Infectious Focus Assay for Determination of Filovirus Titers and Direct Comparison to Plaque and TCID50 Assays

2021 ◽  
Vol 9 (1) ◽  
pp. 156
Author(s):  
Patrick T. Keiser ◽  
Manu Anantpadma ◽  
Hilary Staples ◽  
Ricardo Carrion ◽  
Robert A. Davey
Keyword(s):  
High Throughput Screening ◽  
Plaque Assay ◽  
Neutral Red ◽  
Human Errors ◽  
Subjective Analysis ◽  
Infectious Focus ◽  
Computer Based ◽  
Zaire Ebolavirus ◽  
Focus Assay ◽  
Automated Image Processing

Ongoing efforts to develop effective therapies against filoviruses rely, to different extents, on quantifying the amount of viable virus in samples by plaque, TCID50, and focus assays. Unfortunately, these techniques have inherent variance, and laboratory-specific preferences make direct comparison of data difficult. Additionally, human errors such as operator errors and subjective bias can further compound the differences in outcomes. To overcome these biases, we developed a computer-based automated image-processing method for a focus assay based on the open-source CellProfiler software platform, which enables high-throughput screening of many treatment samples at one time. We compared virus titers calculated using this platform to plaque and TCID50 assays using common stocks of virus for 3 major Filovirus species, Zaire ebolavirus, Sudan ebolavirus, and Marburg marburgvirus with each assay performed by multiple operators on multiple days. We show that plaque assays give comparable findings that differ by less than 3-fold. Focus-forming unit (FFU) and TCID50 assays differ by 10-fold or less from the plaque assays due a higher (FFU) and lower (TCID50) sensitivity. However, reproducibility and accuracy of each assay differs significantly with Neutral Red Agarose Overlay plaque assays and TCID50 with the lowest reproducibility due to subjective analysis and operator error. Both crystal violet methylcellulose overlay plaque assay and focus assays perform best for accuracy and the focus assay performs best for speed and throughput.

Transitioning Nuclear Power Plant Main Control Room From Paper Based Procedures to Computer Based Procedures

2018 ◽  
Vol 62 (1) ◽  
pp. 1605-1609 ◽  
Author(s):  
Roger Lew ◽  
Ronald L. Boring ◽  
Thomas A. Ulrich
Keyword(s):  
Nuclear Power ◽  
Power Plants ◽  
Hybrid Control ◽  
The United States ◽  
Maintenance Cost ◽  
Human Errors ◽  
Nuclear Control ◽  
Computer Based ◽  
Control Rooms ◽  
Analog Systems

The United States (U.S.) has 99 operating Nuclear Power Plants (NPPs). The majority of these were designed and commissioned in the 1970s and 1980s. Plants are modernizing their control systems and main control rooms to be able to continue operating past their original 40-year license agreements. U.S. NPP main control rooms are migrating towards hybrid controls with both digital and analog systems. Digital upgrades, while costly, provide improved reliability, reduced maintenance cost, and the potential for fewer unplanned outages and fewer human errors. U.S. utilities have been slow to embrace computerized procedure system (CBP) research, even though CBPs demonstrate clear operational and human factors benefits. Most of the CBP research has been oriented to new reactor designs or full digital control rooms and is not applicable to the piecemeal modernization approach favored by U.S. plants. Research is needed to examine how CBPs impact operations in hybrid control rooms, and how current paper based procedures can be efficiently migrated to computerized platforms. Work is underway to develop tools and perform the obligatory research needed to design and validate CBPs for modernized U.S. nuclear control rooms.

Transforming Your Robotics Into an Infrastructure for the Future

1997 ◽  
Vol 2 (3) ◽  
pp. 139-143 ◽  
Author(s):  
John Babiak
Keyword(s):  
Data Management ◽  
High Throughput ◽  
High Throughput Screening ◽  
Core Competencies ◽  
The Other ◽  
Exploratory Research ◽  
Screening Process ◽  
Screen Design ◽  
Bar Codes ◽  
Computer Based

The four infrastructural elements for effective high throughput screening are Sample Sourcing, Screen Design, Robotic Hardware, and Data Management. These must all work together to achieve a balance between flexibility and productivity. Sample Sourcing can enhance the high throughput screening process when bar codes are used in a consistent and sensible manner which takes into account the way people think and work. Exploratory research that leads to core competencies in Screen Design can vastly streamline the development and implementation of a high throughput screen by providing scientist customers with specific guidelines which capitalize on your robotic strengths. Robotic Hardware architecture should be open to extension, computer-based, reliable, easy to use, and modified on an ongoing basis. Data Management needs to be pervasive and tie together the other components of the infrastructure. The essential ingredients, though, are motivated and well-trained people who will transform the tools of robotics and automation into the infrastructure needed for effective drug discovery.

scholarly journals Protocol development for discovery of angiogenesis inhibitors via automated methods using zebrafish

2019 ◽  
Author(s):  
Antonio Mauro ◽  
Robin Ng ◽  
Jamie Yuanjun Li ◽  
Rui Guan ◽  
Youdong Wang ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Angiogenesis Inhibitor ◽  
Angiogenesis Inhibitors ◽  
Inhibitory Effect ◽  
Human Errors ◽  
High Fecundity ◽  
Protocol Development ◽  
Compound Libraries ◽  
Automated Methods

AbstractTheir optical clarity as larvae and embryos, small size, and high fecundity make zebrafish ideal for whole animal high throughput screening. A high-throughput drug discovery platform (HTP) has been built to perform fully automated screens of compound libraries with zebrafish embryos. A Tg(Flk1:EGFP) line, marking endothelial cell cytoplasm, was used in this work to help develop protocols and functional algorithms for the system, with the intent of screening for angiogenesis inhibitors. Indirubin 3’ Monoxime (I3M), a known angiogenesis inhibitor, was used at various concentrations to validate the protocols. Consistent with previous studies, a dose dependant inhibitory effect of I3M on angiogenesis was confirmed. The methods and protocols developed here could significantly increase the throughput of drug screens, while limiting human errors. These methods are expected to facilitate the discovery of novel anti-angiogenesis compounds and can be adapted for many other applications in which samples have a good fluorescent signal.

scholarly journals Rapid Quantification and Neutralization Assays for Novel Coronavirus SARS-CoV-2 Using Avicel RC-591 Semi-Solid Overlay

2020 ◽  
Author(s):  
Anna N. Honko ◽  
Nadia Storm ◽  
David J. Bean ◽  
Jhonatan Henao Vasquez ◽  
Sierra N. Downs ◽  
...  
Keyword(s):  
Plaque Assay ◽  
Neutral Red ◽  
The Novel ◽  
Invaluable Tool ◽  
Automated Processing ◽  
Pandemic Response ◽  
Semi Solid ◽  
Novel Coronavirus ◽  
Pathogenic Agents ◽  
Potential Therapeutics

When working with the novel coronavirus SARS-CoV-2 during a pandemic response, having a rapid, reproducible and reliable assay for infectious virus quantitation and utilization for evaluation of potential therapeutics is critical. Compared to traditional agarose overlay plaques visualized with neutral red, assays performed with Avicel R RC-591 semi-solid overlay provide a simplified format for rapid and easy detection and neutralization testing. The method is easily modified for higher throughput using dispensers or automated processing. Fixation using formalin provides flexibility when dealing with pathogenic agents such as SARS-CoV-2 where tissue culture plates might be removed from biocontainment for staining. Although plaque assays are considered straightforward in principle, having an easily reproducible, consistent plaque assay is an invaluable tool.

scholarly journals Opportunities and Challenges of Online Take-Home Exams in Medical Education

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Abtin Heidarzadeh ◽  
Hanieh Zehtab Hashemi ◽  
Parvane Parvasideh ◽  
Zahra Hasan Larijani ◽  
Parisa Baqhdadi ◽  
...  
Keyword(s):  
Higher Education ◽  
Student Assessment ◽  
Web Pages ◽  
Medical Sciences ◽  
Human Errors ◽  
Unethical Conduct ◽  
Examination Method ◽  
General Search ◽  
Computer Based ◽  
Ongoing Monitoring

Context: Student assessment is an essential part of higher education. Many different technology-based assessment methods have been formed with the increasing development of IT and its introduction into the education system. Online take-home exams are computer-based exams in which the examinees can take at a place of their own choice and on their own computers. Despite its benefits, this method is faced with certain problems. The present study investigates the challenges in holding take-home computer-based exams in medical sciences and various solutions proposed to use this method more extensively in Iran in situations of crisis. Evidence Acquisition: The present review article was drafted upon a search conducted in Scopus, Google Scholar, and Google’s general search engine using the following keywords and search strategies: "Take-home exam", OR "Take-home assessment", OR, "Online exam", OR "Online assessment", AND "Higher education". The content of the related documents published from 2009 to 2020, including articles, books, and web pages, was selected and assessed, and 35 articles were finally used to accomplish the study objectives. Results: Online take-home exams have many advantages, including reduced human errors, rapid scoring, and reduced stress on the examinees. Nonetheless, one of the limitations of this examination method is that the examinees may not meet all the criteria required for taking exams at home. The obvious risk is students’ unethical conduct and cheating, which composes a major challenge of this examination modality. Conclusions: The reliability and correctness of exams can be improved using combination techniques, question banks, and giving random equivalent questions to each candidate that are not necessarily similar, and also mixing up the questions and their answers, which can provide a tool for preventing or limiting cheating. Online monitoring systems are also one of the strategies proposed for ongoing monitoring of online exams by an invigilator that are generally developed through artificial intelligence.

Recording and Evaluation of Human Factor Events with a View to System Awareness and Ergonomic Weak Points within the System, at the Example of Commercial Aeronautics

2000 ◽  
Vol 44 (22) ◽  
pp. 835-838 ◽  
Author(s):  
Bernd Linsenmaier ◽  
Oliver Straeter
Keyword(s):  
Situation Awareness ◽  
Human Factor ◽  
Point Of View ◽  
Sufficient Information ◽  
Event Analysis ◽  
Human Errors ◽  
Analysis Scheme ◽  
Weak Points ◽  
Computer Based ◽  
Almost All

The risk for human errors is particularly high, if a person has not considered enough information about his situation. This consideration of information also can be described as situation awareness. It will become dangerous, if a person assesses his awareness higher as it is actual. This discrepancy between subjective and actual situation awareness can be found out at almost all Human Factor (HF) Events. With the examination of the situation awareness, the event analysis is carried out consistently from the point of view of the man. By this concept, HF Events can be examined even more on ergonomic aspects. It is however necessary that reports about HF Events provide sufficient information about the situation awareness. For this a computer-based event analysis scheme is used to report events interactively. With this software, we currently investigate how it is possible to create uniform and comparable event reports. The complete event is divided up into sub-events and is described using the Man Machine System framework. The demarcation of the sub-events is mainly made by allocation of the persons involved. This human and system related view also allows describing the situation awareness of this person and provides data for the later analysis of the situation awareness.

scholarly journals High-throughput Screening and Experimental Identification of Potent Drugs Targeting SARS-CoV-2 Main Protease

2020 ◽  
Author(s):  
Yan Li ◽  
Jinyong Zhang ◽  
Ning Wang ◽  
Yanjing Zhang ◽  
Yongjun Yang ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Drug Targets ◽  
Binding Pocket ◽  
Substrate Binding Pocket ◽  
Stable Conformation ◽  
Cross Docking ◽  
Main Protease ◽  
Computer Based ◽  
Old Drugs

Abstract The main protease (Mpro) is one of the best-characterized drug targets among coronaviruses. In the current study, we adopted a multiple cross-docking strategy against different crystal structures of SARS-CoV-2 Mpro to perform computer-based high-throughput virtual screening of possible inhibitors from a drug database using Autodock Vina and SeeSAR software, combined with our in-house automatic processing scripts. The KDs between screened candidates and Mpro were determined using Biacore. Seven drugs were found to fit the substrate-binding pocket of Mpro with a stable conformation, showing high KDs that ranged from 6.79E-7 M to 5.20E-5 M. Finally, mutagenesis studies confirmed that these drugs interact with Mpro specifically, suggesting that our method was reliable and convincing. Given the safety of these old drugs, they may serve as promising candidates to treat the infection of SARS-CoV-2. Our results also provide rational explanations for the behaviour of five drugs evaluated in clinical trials.

scholarly journals Plaque assay and improved yield of human coronaviruses in a human rhabdomyosarcoma cell line

1979 ◽  
Vol 9 (6) ◽  
pp. 722-728 ◽  
Author(s):  
O W Schmidt ◽  
M K Cooney ◽  
G E Kenny
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Maximum Yield ◽  
Plaque Assay ◽  
Growth Curves ◽  
Neutral Red ◽  
Virus Propagation ◽  
Extracellular Virus ◽  
Rd Cells ◽  
Intracellular Virus

Propagation and plaque assay of human coronavirus prototypes were studied in two human cell lines: a diploid fetal tonsil (FT) and a heteroploid rhabdomyosarcoma (RD) cell lines. Plaques, observed within 2 to 3 days on FT cell monolayers with both 229E and OC43 viruses, appeared as colorless areas after staining with neutral red or crystal violet, whereas neutral red staining was required for visualization of plaques on RD cells. The plating efficiencies were approximately equal between the two cell lines, but virus assay by plaque formation was 15- to 30-fold more efficient than tube dilution assay with 50% endpoints. The discrepancy between 50% endpoint and plaque-forming unit values was striking and appeared to result from the fact that killing of cells (particularly RD cells) by coronaviruses was not accompanied by visible changes in the cells but killing was detected by the failure of infected cells to stain with a vital dye. The latent phase in one-step growth curves was 5 to 6 h for both viruses in either cell line, but the maximum yield of intracellular virus was reached in 18 to 20 h for FT cells and 24 to 28 h for RD cells. Virus release also differed between the two cell lines: in FT cells, the maximum yield of extracellular virus was reached 2 to 3 h later than that of intracellular virus, whereas in RD cells, the difference was 5 h for 229E virus and 10 h for OC43 virus. Although both cell lines appear equally useful for plaque assay, RD cells would be preferred for mass virus propagation because yields (5 X 10(8) plaque-forming units per ml) were 10-fold higher than in FT cells, a finding true for both virus prototypes.

Plaque reduction neutralization test for human cytomegalovirus based upon enhanced uptake of neutral red by virus-infected cells

1976 ◽  
Vol 4 (1) ◽  
pp. 61-66
Author(s):  
N J Schmidt ◽  
J Dennis ◽  
E H Lennette
Keyword(s):  
Human Cytomegalovirus ◽  
Neutralization Test ◽  
Plaque Assay ◽  
Neutralizing Antibody ◽  
Neutral Red ◽  
Plaque Reduction Neutralization Test ◽  
Cell Monolayers ◽  
Human Sera ◽  
Antibody Titers ◽  
Infected Cells

Foci of cells infected with human cytomegalovirus were noted to stain more intensely than uninfected cells with neutral red, and this provided the basis for development of a plaque assay and plaque reduction neutralization test for cytomegalovirus. Plaques demonstrable by neutral red staining could be counted at 8 days after infection; thus, results could be obtained earlier than for plaque assay systems based upon the viral cytopathic effect, a fewer manipulations were required for staining cell monolayers to demonstrate plaques. Certain variables affecting plaque size and numbers and antibody titers were defined. Addition of fresh guinea pig complement to the reaction mixtures markedly enhanced cytomegalovirus-neutralizing antibody titers of hyperimmune animal sera, but titers of human sera were enhanced only two-or fourfold.

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