Dataset Information

Evaluation of the automated MicroFlow^® and Metafer™ platforms for high-throughput micronucleus scoring and dose response analysis in human lymphoblastoid TK6 cells.

ABSTRACT: The use of manual microscopy for the scoring of chromosome damage in the in vitro micronucleus assay is often associated with user subjectivity. This level of subjectivity can be reduced by using automated platforms, which have added value of faster with high-throughput and multi-endpoint capabilities. However, there is a need to assess the reproducibility and sensitivity of these automated platforms compared with the gold standard of the manual scoring. The automated flow cytometry-based MicroFlow^® and image analysis-based Metafer™ were used for dose response analyses in human lymphoblastoid TK6 cells exposed to the model clastogen, methyl methanesulfonate (MMS), aneugen, carbendazim, and the weak genotoxic carcinogen, ochratoxin A (OTA). Cells were treated for 4 or 30 h, with a 26- or 0-h recovery. Flow cytometry scoring parameters and the Metafer™ image classifier were investigated, to assess any potential differences in the micronucleus (MN) dose responses. Dose response data were assessed using the benchmark dose approach with chemical and scoring system set as covariate to assess reproducibility between endpoints. A clear increase in MN frequency was observed using the MicroFlow^® approach on TK6 cells treated for 30 h with MMS, carbendazim and OTA. The MicroFlow^®-based MN frequencies were comparable to those derived by using the Metafer™ and manual scoring platforms. However, there was a potential overscoring of MN with the MicroFlow^® due to the cell lysis step and an underscoring with the Metafer™ system based on current image classifier settings. The findings clearly demonstrate that the MicroFlow^® and Metafer™ MN scoring platforms are powerful tools for automated high-throughput MN scoring and dose response analysis.

SUBMITTER: Verma JR

PROVIDER: S-EPMC5489585 | biostudies-literature | 2017 Jul

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Evaluation of the automated MicroFlow<sup>®</sup> and Metafer™ platforms for high-throughput micronucleus scoring and dose response analysis in human lymphoblastoid TK6 cells.

Verma Jatin R JR Rees Benjamin J BJ Wilde Eleanor C EC Thornton Catherine A CA Jenkins Gareth J S GJS Doak Shareen H SH Johnson George E GE

Archives of toxicology 20161210 7

The use of manual microscopy for the scoring of chromosome damage in the in vitro micronucleus assay is often associated with user subjectivity. This level of subjectivity can be reduced by using automated platforms, which have added value of faster with high-throughput and multi-endpoint capabilities. However, there is a need to assess the reproducibility and sensitivity of these automated platforms compared with the gold standard of the manual scoring. The automated flow cytometry-based MicroF ...[more]

PMID: 27942789

Similar Datasets

Project description:Introduction: Higher-throughput mode-of-action-based assays provide a valuable approach to expedite chemical evaluation for human health risk assessment. In this study, we combined the high-throughput alkaline CometChip® assay with the TGx-DDI transcriptomic biomarker (DDI = DNA damage-inducing) using high-throughput TempO-Seq® as an integrated genotoxicity testing approach. Objective: To determine if the high-throughput CometChip® assay and TGx-DDI biomarker analysis can be combined using human HepaRG™ cells to provide an efficient next-generation genotoxicity screening approach to identify DNA damage-inducing (DDI) chemicals. Methods: Metabolically competent human HepaRG™ cell cultures were exposed to increasing concentrations of 12 chemicals (9 DDI and 3 non-DDI) using a repeat exposure design (0 hr, 24 hr, 48 hr) in 96-well plates to measure and classify chemicals based on their ability to damage DNA. After a 4 hr recovery period, exposed cells were used to perform the Trevigen CometChip assay to assess DNA damage or used for TempO-Seq® S1500+ Targeted Transcriptome Sequencing for TGx-DDI classification purposes. Bioinformatics and statistical tools were used to classify chemicals as DDI or non-DDI using three separate analyses. Benchmark concentration analysis was also conducted for both assays for the purposes of potency ranking. Results: In combination, the CometChip® assay and the TGx-DDI were 100% accurate in identifying chemicals that induce DNA damage. Quantitative benchmark concentration (BMC) modeling was applied to evaluate chemical potencies for both assays. The BMCs for the CometChip® assay and the TGx-DDI for all 12 chemicals were highly concordant (within four-fold) and resulted in identical potency rankings. Conclusions: These results suggest that these two assays can be integrated for efficient, high-throughput identification of DNA damage in HepaRG™ cells. This study provides evidence for the complementarity of the high-throughput CometChip® assay with TGx-DDI biomarker analysis. Integration of these two tests provides an effective and high-throughput approach to identify DDI agents. This work is a first step in accomplishing a more integrated genotoxicity testing strategy to derive mechanistic information to better inform human health risk assessment in a high-throughput manner.

Project description:BackgroundThe use of SharePoint® collaboration software for content management has become a critical part of today's drug discovery process. SharePoint 2010 software has laid a foundation which enables researchers to collaborate and search on various contents. The amount of data generated during a transition of a single compound from preclinical discovery to commercialization can easily range in terabytes, thus there is a greater demand of a chemically aware search algorithm that supplements SharePoint which enables researchers to query for information in a more intuitive and effective way. Thus by supplementing SharePoint with Chemically Aware™ features provides a great value to the pharmaceutical and biotech companies and makes drug discovery more efficient. Using several tools we have integrated SharePoint with chemical, compound, and reaction databases, thereby improving the traditional search engine capability and enhancing the user experience.ResultsThis paper describes the implementation of a Chemically Aware™ system to supplement SharePoint. A Chemically Aware SharePoint (CASP) allows users to tag documents by drawing a structure and associating it with the related content. It also allows the user to search SharePoint software content and internal/external databases by carrying out substructure, similarity, SMILES, and IUPAC name searches. Building on traditional search, CASP takes SharePoint one step further by providing a intuitive GUI to the researchers to base their search on their knowledge of chemistry than textual search. CASP also provides a way to integrate with other systems, for example a researcher can perform a sub-structure search on pdf documents with embedded molecular entities.ConclusionA Chemically Aware™ system supplementing SharePoint is a step towards making drug discovery process more efficient and also helps researchers to search for information in a more intuitive way. It also helps the researchers to find information which was once difficult to find by allowing one to tag documents with molecular entities and integrating with image recognition software to find information from pdf documents.

Project description:Automated methods are needed to facilitate high-throughput and reproducible scoring of Ki67 and other markers in breast cancer tissue microarrays (TMAs) in large-scale studies. To address this need, we developed an automated protocol for Ki67 scoring and evaluated its performance in studies from the Breast Cancer Association Consortium. We utilized 166 TMAs containing 16,953 tumour cores representing 9,059 breast cancer cases, from 13 studies, with information on other clinical and pathological characteristics. TMAs were stained for Ki67 using standard immunohistochemical procedures, and scanned and digitized using the Ariol system. An automated algorithm was developed for the scoring of Ki67, and scores were compared to computer assisted visual (CAV) scores in a subset of 15 TMAs in a training set. We also assessed the correlation between automated Ki67 scores and other clinical and pathological characteristics. Overall, we observed good discriminatory accuracy (AUC = 85%) and good agreement (kappa = 0.64) between the automated and CAV scoring methods in the training set. The performance of the automated method varied by TMA (kappa range= 0.37-0.87) and study (kappa range = 0.39-0.69). The automated method performed better in satisfactory cores (kappa = 0.68) than suboptimal (kappa = 0.51) cores (p-value for comparison = 0.005); and among cores with higher total nuclei counted by the machine (4,000-4,500 cells: kappa = 0.78) than those with lower counts (50-500 cells: kappa = 0.41; p-value = 0.010). Among the 9,059 cases in this study, the correlations between automated Ki67 and clinical and pathological characteristics were found to be in the expected directions. Our findings indicate that automated scoring of Ki67 can be an efficient method to obtain good quality data across large numbers of TMAs from multicentre studies. However, robust algorithm development and rigorous pre- and post-analytical quality control procedures are necessary in order to ensure satisfactory performance.

Dataset Information

Evaluation of the automated MicroFlow^® and Metafer™ platforms for high-throughput micronucleus scoring and dose response analysis in human lymphoblastoid TK6 cells.

Publications

Evaluation of the automated MicroFlow<sup>®</sup> and Metafer™ platforms for high-throughput micronucleus scoring and dose response analysis in human lymphoblastoid TK6 cells.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets

Dataset Information

Evaluation of the automated MicroFlow® and Metafer™ platforms for high-throughput micronucleus scoring and dose response analysis in human lymphoblastoid TK6 cells.

Publications

Evaluation of the automated MicroFlow<sup>®</sup> and Metafer™ platforms for high-throughput micronucleus scoring and dose response analysis in human lymphoblastoid TK6 cells.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets

Evaluation of the automated MicroFlow^® and Metafer™ platforms for high-throughput micronucleus scoring and dose response analysis in human lymphoblastoid TK6 cells.