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60-539: Originally titled the Chemistry Dashboard, the first version was released in 2016. The latest release of the database (version 3.0.5) contains manually curated data for over 875,000 chemicals and incorporates the latest data generated from the EPA's Toxicity Forecaster (ToxCast) high-throughput screening program. The Chemicals Dashboard incorporates data from several previous EPA databases into one package including

120-646: A de facto standard height of 44 mm. Microplates used for PCR are designed to have a notably thinner wall thickness than standard ANSI/SLAS microplates (to allow for better thermal conduction ), and to come in a few different "skirt" types: full-skirt, half-skirt or semi-skirted, and unskirted or no-skirted. The skirt is analogous to the footprint & flange of the ANSI/SLAS standards, so while most full-skirt PCR microplates may be ANSI/SLAS compliant, other deviations such as semi-skirted or others, are not compliant ANSI/SLAS standards. The earliest microplate

180-415: A ResonantAcoustic mixer, Merck reported reduced processing time to less than 2 hours on only 1-2 mg of drug compound per well. Merck also indicated the acoustic milling approach allows for the preparation of high dose nanosuspension formulations that could not be obtained using conventional milling equipment. Whereby traditional HTS drug discovery uses purified proteins or intact cells, recent development of

240-442: A consequence, robust methods such as the z*-score method, SSMD*, B-score method, and quantile-based method have been proposed and adopted for hit selection. In a screen with replicates, we can directly estimate variability for each compound; as a consequence, we should use SSMD or t-statistic that does not rely on the strong assumption that the z-score and z*-score rely on. One issue with the use of t-statistic and associated p-values

300-574: A corner notch (aka chamfer) is shown at the A1 (top-left) corner in the ANSI SLAS drawings, and many microplates do implement this A1 corner notch, in actuality the "quantity and location of chamfers(s) is optional", so in practice the presence or absence of corner notches at additional corners (i.e. the bottom-left) is a proprietary implementation which causes difficulties with accessory cross-compatibility such as with microplate lids that may also implement

360-584: A desired size of effects in an HTS is called a hit. The process of selecting hits is called hit selection. The analytic methods for hit selection in screens without replicates (usually in primary screens) differ from those with replicates (usually in confirmatory screens). For example, the z-score method is suitable for screens without replicates whereas the t-statistic is suitable for screens with replicates. The calculation of SSMD for screens without replicates also differs from that for screens with replicates . For hit selection in primary screens without replicates,

420-574: A finished product. ELISA plates may now be assembled from twelve separate strips of eight wells, making it easier to only partially use a plate. Microplates are produced with the same standardized footprint, but using a variety of formats (see table below), materials (see above section ), plate heights, numbers of wells, well shapes, and well bottom heights, with some of these characteristics being more varied between manufacturers than others (see below section ). There are also less common 192- and 768-well plates. An attempt at standardizing microplates

480-679: A gatekeeper for excellent quality assays. In a typical HTS experiment, a clear distinction between a positive control and a negative reference such as a negative control is an index for good quality. Many quality-assessment measures have been proposed to measure the degree of differentiation between a positive control and a negative reference. Signal-to-background ratio, signal-to-noise ratio, signal window, assay variability ratio, and Z-factor have been adopted to evaluate data quality. Strictly standardized mean difference ( SSMD ) has recently been proposed for assessing data quality in HTS assays. A compound with

540-418: A library of stock plates , whose contents are carefully catalogued, and each of which may have been created by the lab or obtained from a commercial source. These stock plates themselves are not directly used in experiments; instead, separate assay plates are created as needed. An assay plate is simply a copy of a stock plate, created by pipetting a small amount of liquid (often measured in nanoliters ) from

600-667: A library of over 380,000 compounds. Northwestern University's High Throughput Analysis Laboratory supports target identification, validation, assay development, and compound screening. The non-profit Sanford Burnham Prebys Medical Discovery Institute also has a long-standing HTS facility in the Conrad Prebys Center for Chemical Genomics which was part of the MLPCN. The non-profit Scripps Research Molecular Screening Center (SRMSC) continues to serve academia across institutes post-MLPCN era. The SRMSC uHTS facility maintains one of

660-418: A microplate typically holds somewhere between tens of nanolitres to several millilitres of liquid. They can also be used to store dry powder or as racks to support glass tube inserts. Wells can be either circular or square. For compound storage applications, square wells with close fitting silicone cap-mats are preferred. Microplates can be stored at low temperatures for long periods, may be heated to increase

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720-423: A more physiologically relevant format. HTS is a relatively recent innovation, made feasible largely through modern advances in robotics and high-speed computer technology. It still takes a highly specialized and expensive screening lab to run an HTS operation, so in many cases a small- to moderate-size research institution will use the services of an existing HTS facility rather than set up one for itself. There

780-493: A number of experiments on the wells (such as shining polarized light on them and measuring reflectivity, which can be an indication of protein binding). In this case, the machine outputs the result of each experiment as a grid of numeric values, with each number mapping to the value obtained from a single well. A high-capacity analysis machine can measure dozens of plates in the space of a few minutes like this, generating thousands of experimental datapoints very quickly. Depending on

840-506: A particular biomolecular pathway. The results of these experiments provide starting points for drug design and for understanding the noninteraction or role of a particular location. The key labware or testing vessel of HTS is the microtiter plate , which is a small container, usually disposable and made of plastic, that features a grid of small, open divots called wells . In general, microplates for HTS have either 96, 192, 384, 1536, 3456 or 6144 wells. These are all multiples of 96, reflecting

900-427: A preferred value or limits for those dimensional definitions. Therefore all well bottom heights are currently proprietary implementations without a clear de facto standard. This lack of standardization can cause difficulties with applications such as automated autosampler needle injection. The height of a standard microplate is also defined, however this is sometimes not followed by manufacturers, even if they follow

960-463: A scientist does not understand some statistics or rudimentary data-handling technologies, he or she may not be considered to be a true molecular biologist and, thus, will simply become "a dinosaur." High-quality HTS assays are critical in HTS experiments. The development of high-quality HTS assays requires the integration of both experimental and computational approaches for quality control (QC). Three important means of QC are (i) good plate design, (ii)

1020-403: A silicon sheet of lenses that can be placed over microfluidic arrays to allow the fluorescence measurement of 64 different output channels simultaneously with a single camera. This process can analyze 200,000 drops per second. In 2013, researchers have disclosed an approach with small molecules from plants. In general, it is essential to provide high-quality proof-of-concept validations early in

1080-836: A single construct such as a single siRNA or cDNA. Functional genomics is typically paired with high content screening using e.g. epifluorescent microscopy or laser scanning cytometry. The University of Illinois also has a facility for HTS, as does the University of Minnesota. The Life Sciences Institute at the University of Michigan houses the HTS facility in the Center for Chemical Genomics. Columbia University has an HTS shared resource facility with ~300,000 diverse small molecules and ~10,000 known bioactive compounds available for biochemical, cell-based and NGS-based screening. The Rockefeller University has an open-access HTS Resource Center HTSRC (The Rockefeller University, HTSRC ), which offers

1140-731: A specific research topic (such as additives in cigarettes or chemicals demonstrating effects on neurodevelopmental effects ) or the specific assay endpoints they are covered by. Within the online dashboard searches can be performed by product/use categories, assay/gene, systematic name, synonym, CAS number , DSSTox Substance ID or InChiKey . Under the Advanced Search tab chemicals can be searched based on their mass or molecular formula . Searches can also be performed for groups of chemicals based on Chemical Name CASRN, InChIKey, DSSTox Substance ID, DSSTox Compound ID, InChIKey Skeleton, MS-Ready Formula, Exact Formula, or Monoisotopic Mass using

1200-581: A standard definition of a microplate. A series of standards was proposed in 2003 and published by the American National Standards Institute (ANSI) on behalf of the SBS. The standards govern various characteristics of a microplate including well positioning (but not shape, depth, and diameter) as well as plate properties, which allows interoperability between microplates, instrumentation and equipment from different suppliers, and

1260-472: A very challenging drug target. Hits are confirmed at the screening step due to the bell-shaped curve. This method is very similar to the quantitative HTS method (screening and hit confirmation at the same time), except that using this approach greatly decreases the data point number and can screen easily more than 100.000 biological relevant compounds. Switching from an orbital shaker, which required milling times of 24 hours and at least 10 mg of drug compound to

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1320-510: A web-based version of the "Abstract Sifter", Google Scholar and reports from EPA's Provisional Peer Reviewed Toxicity Values ( PPRTV ) and the EPA Integrated Risk Information System ( IRIS ). Real-time QSAR prediction for multiple physicochemical property and toxicity endpoints is available through the predictions tab . The dashboard provides support for mass spectrometry providing searches against

1380-558: Is injection molding , using materials such as polystyrene, polypropylene and cyclo-olefin for different temperature and chemical resistance needs. Glass is also a possible material, and vacuum forming can be used with many other plastics such as polycarbonate. Microplates are manufactured from a variety of materials: Composite microplates, including filter bottom plates, solid phase extraction (SPE) plates, and even some advanced PCR plate designs, use multiple components and/or materials which are moulded separately and later assembled into

1440-477: Is a method for scientific discovery especially used in drug discovery and relevant to the fields of biology , materials science and chemistry . Using robotics , data processing/control software, liquid handling devices, and sensitive detectors, high-throughput screening allows a researcher to quickly conduct millions of chemical, genetic, or pharmacological tests. Through this process one can quickly recognize active compounds, antibodies, or genes that modulate

1500-440: Is a trend in academia for universities to be their own drug discovery enterprise. These facilities, which normally are found only in industry, are now increasingly found at universities as well. UCLA , for example, features an open access HTS laboratory Molecular Screening Shared Resources (MSSR, UCLA), which can screen more than 100,000 compounds a day on a routine basis. The open access policy ensures that researchers from all over

1560-403: Is comparable across experiments and, thus, we can use the same cutoff for the population value of SSMD to measure the size of compound effects . Unique distributions of compounds across one or many plates can be employed either to increase the number of assays per plate or to reduce the variance of assay results, or both. The simplifying assumption made in this approach is that any N compounds in

1620-438: Is tested across four- to five-orders of magnitude of concentrations. Microtiter plate A microplate , also known as a microtiter plate , microwell plate or multiwell , is a flat plate with multiple "wells" used as small test tubes. The microplate has become a standard tool in analytical research and clinical diagnostic testing laboratories. A very common usage is in the enzyme-linked immunosorbent assay (ELISA),

1680-411: Is that they are affected by both sample size and effect size. They come from testing for no mean difference, and thus are not designed to measure the size of compound effects. For hit selection, the major interest is the size of effect in a tested compound. SSMD directly assesses the size of effects. SSMD has also been shown to be better than other commonly used effect sizes. The population value of SSMD

1740-413: Is the most likely agent, while also providing three measurements of compound B's efficacy against the specified target. Commercial applications of this approach involve combinations in which no two compounds ever share more than one well, to reduce the (second-order) possibility of interference between pairs of compounds being screened. Automation and low volume assay formats were leveraged by scientists at

1800-508: The NIH Chemical Genomics Center (NCGC) to develop quantitative HTS (qHTS), a paradigm to pharmacologically profile large chemical libraries through the generation of full concentration-response relationships for each compound. With accompanying curve fitting and cheminformatics software qHTS data yields half maximal effective concentration (EC50), maximal response, Hill coefficient (nH) for the entire library enabling

1860-630: The ToxCast Dashboard, the Endocrine Disruption Screening Program (EDSP) Dashboard and the Chemical and Products Database (CPDat). The CompTox Chemicals Dashboard database contains high quality chemical structures and information that have been extensively curated and quality checked, which can be used as a resource for analytical scientists involved in structure identification. Chemical hazard data in

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1920-502: The assessment of nascent structure activity relationships (SAR). In March 2010, research was published demonstrating an HTS process allowing 1,000 times faster screening (100 million reactions in 10 hours) at 1-millionth the cost (using 10 times the reagent volume) than conventional techniques using drop-based microfluidics. Drops of fluid separated by oil replace microplate wells and allow analysis and hit sorting while reagents are flowing through channels. In 2010, researchers developed

1980-402: The basis of most modern medical diagnostic testing in humans and animals. A microplate typically has 6, 12, 24, 48, 96, 384 or 1536 sample wells arranged in a 2:3 rectangular matrix . Some microplates have been manufactured with 3456 or 9600 wells, and an "array tape" product has been developed that provides a continuous strip of microplates embossed on a flexible plastic tape. Each well of

2040-410: The batch search function. An automated read-across tool called Generalized Read-Across (GenRA) is integrated into The Chemicals Dashboard. GenRA is designed to keep the expert consideration inherent in the read-across method, but automate the chemical selection process to help predict toxicity. The Dashboard also has the capability to search existing scientific literature sources such as PubMed , via

2100-633: The chemical data contained in the database based on mass and molecular formula . The dashboard has been applied to non-targeted analysis searching for " known unknowns ". Both targeted mass spectrometry and non-targeted mass spectrometry are supported. The searches utilize a search based on "MS-Ready" forms of chemical compounds. Individual chemical substances are collapsed into a form that would be detected by mass spectrometry such that salts are desalted and neutralized and multi-component chemicals are separated into their individual components. High-throughput screening High-throughput screening ( HTS )

2160-427: The compounds in the wells, measurements are taken across all the plate's wells, either manually or by a machine. Manual measurements are often necessary when the researcher is using microscopy to (for example) seek changes or defects in embryonic development caused by the wells' compounds, looking for effects that a computer could not easily determine by itself. Otherwise, a specialized automated analysis machine can run

2220-634: The dashboard comes from both traditional laboratory animal studies and high-throughput screening . Biological data from high-throughput screening is generated by EPA's ToxCast program, the ToxCast data in the database provides information about the assays used and their response potency and efficacy. These data can be found in the bioactivity tab. The Chemicals Dashboard can be accessed via a web interface or sets of data within it can be downloaded for use offline. The Lists tab can be used to browse and download groups of related chemicals based on their relevance to

2280-493: The data-collection process. HTS robots that can test up to 100,000 compounds per day currently exist. Automatic colony pickers pick thousands of microbial colonies for high throughput genetic screening. The term uHTS or ultra-high-throughput screening refers (circa 2008) to screening in excess of 100,000 compounds per day. With the ability of rapid screening of diverse compounds (such as small molecules or siRNAs ) to identify active compounds, HTS has led to an explosion in

2340-415: The drug discovery process. Here technologies that enable the identification of potent, selective, and bioavailable chemical probes are of crucial interest, even if the resulting compounds require further optimization for development into a pharmaceutical product. Nuclear receptor RORα, a protein that has been targeted for more than a decade to identify potent and bioavailable agonists, was used as an example of

2400-473: The easily interpretable ones are average fold change, mean difference, percent inhibition, and percent activity. However, they do not capture data variability effectively. The z-score method or SSMD, which can capture data variability based on an assumption that every compound has the same variability as a negative reference in the screens. However, outliers are common in HTS experiments, and methods such as z-score are sensitive to outliers and can be problematic. As

2460-601: The footprint and flange standards. There are also deep well microplates sometimes called "blocks". Unlike plates of normal height, the ANSI SLAS 2-2004 height standard, does not define a standard height for deep well plates (blocks). Deepwell plates do typically follow a de facto standard height of 44 mm. Reservoir plates are also commercially available. Reservoir plates have columns of wells (as in 96-well, 24-well, etc. plates) that are fused into single wells, so that they provide additional volume for multichannel pipettes. Like deepwell plates or blocks, they often follow

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2520-452: The function of each gene in the context of interest by either knocking each gene out or overexpressing it. Parallel access to high-throughput small molecule screen and a genome wide screen enables researchers to perform target identification and validation for given disease or the mode of action determination on a small molecule. The most accurate results can be obtained by use of "arrayed" functional genomics libraries, i.e. each library contains

2580-645: The largest library collections in academia, presently at well-over 665,000 small molecule entities, and routinely screens the full collection or sub-libraries in support of multi-PI grant initiatives. In the United States, the National Institutes of Health or NIH has created a nationwide consortium of small-molecule screening centers to produce innovative chemical tools for use in biological research. The Molecular Libraries Probe Production Centers Network, or MLPCN, performs HTS on assays provided by

2640-492: The matching corner notch. The well position is also standardized, but only for 96- , 384-, and 1536-well plates. These are generally well followed by manufacturers: 96-well plates have a 9 mm well-to-well spacing, 384-wells a 4.5 mm spacing, and 1536-wells a 2.25 mm spacing. A notable characteristic is that the well array is symmetrical when the plate is rotated 180˚ around its Z-axis (height axis). Therefore, scientific instruments which use microplates, can accept

2700-517: The microplate are available in different shapes: Wells also have different geometries at the bottom of the well: Round wells in particular often come in a few diameters: The most recent addition to the ANSI SLAS microplate standards was the inclusion of a well bottom standard. The standard however specifies definitions and test methods only, for the "Microplate Well Bottom Elevation (WBE)", "Well Bottom Elevation Variation (WBEV)", and "Intra-Well Bottom Elevation Variation (IWBEV)", but it does not state

2760-503: The original 96-well microplate with spaced wells of 8 x 12 with 9 mm spacing. Most of the wells contain test items, depending on the nature of the experiment. These could be different chemical compounds dissolved e.g. in an aqueous solution of dimethyl sulfoxide (DMSO). The wells could also contain cells or enzymes of some type. (The other wells may be empty or contain pure solvent or untreated samples, intended for use as experimental controls .) A screening facility typically holds

2820-549: The plate in one of two rotated orientations - either "correct" or 180˚ rotated. Other variants like 24-well plates, are not considered in the standard, but there is a de facto standard to implement to 24-wells by apply the same scaling factor as the 384- to 96-well transition, i.e. 24-wells have an 18 mm spacing. Notably, the shape and diameter of the well is not standardized, and has several proprietary implementations. This causes difficulties with accessory cross-compatibility such as with microplate cap mats . Wells within

2880-475: The rate of data generated in recent years . Consequently, one of the most fundamental challenges in HTS experiments is to glean biochemical significance from mounds of data, which relies on the development and adoption of appropriate experimental designs and analytic methods for both quality control and hit selection . HTS research is one of the fields that have a feature described by John Blume, Chief Science Officer for Applied Proteomics, Inc., as follows: Soon, if

2940-792: The rate of solvent evaporation from their wells and can even be heat-sealed with foil or clear film. Microplates with an embedded layer of filter material were developed in the early 1980s by several companies, and today, there are microplates for just about every application in life science research which involves filtration, separation, optical detection, storage, reaction mixing, cell culture and detection of antimicrobial activity. The enormous growth in studies of whole live cells has led to an entirely new range of microplate products which are " tissue culture treated" especially for this work. The surfaces of these products are modified using an oxygen plasma discharge to make their surfaces more hydrophilic so that it becomes easier for adherent cells to grow on

3000-712: The research community, against a large library of small molecules maintained in a central molecule repository. In addition, the NIH created the National Center for Advancing Translational Sciences or NCATS, housed in Shady Grove Maryland, that carries out small molecule and RNAi screens in collaboration with academic laboratories. Of note, the small molecule screening uses 1536 well plates, a capability rarely seen in academic screening laboratories that allows one to carry out quantitative HTS in which each compound

3060-712: The results of this first assay, the researcher can perform follow up assays within the same screen by "cherrypicking" liquid from the source wells that gave interesting results (known as "hits") into new assay plates, and then re-running the experiment to collect further data on this narrowed set, confirming and refining observations. Automation is an essential element in HTS's usefulness. Typically, an integrated robot system consisting of one or more robots transports assay-microplates from station to station for sample and reagent addition, mixing, incubation, and finally readout or detection. An HTS system can usually prepare, incubate, and analyze many plates simultaneously, further speeding

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3120-405: The same well will not typically interact with each other, or the assay target, in a manner that fundamentally changes the ability of the assay to detect true hits. For example, imagine a plate wherein compound A is in wells 1–2–3, compound B is in wells 2–3–4, and compound C is in wells 3–4–5. In an assay of this plate against a given target, a hit in wells 2, 3, and 4 would indicate that compound B

3180-500: The selection of effective positive and negative chemical/biological controls, and (iii) the development of effective QC metrics to measure the degree of differentiation so that assays with inferior data quality can be identified. A good plate design helps to identify systematic errors (especially those linked with well position) and determine what normalization should be used to remove/reduce the impact of systematic errors on both QC and hit selection. Effective analytic QC methods serve as

3240-976: The surface which would otherwise be strongly hydrophobic . A number of companies have developed robots to specifically handle microplates. These robots may be liquid handlers which aspirate or dispense liquid samples from and to these plates, or "plate movers" which transport them between instruments, plate stackers which store microplates during these processes, plate hotels for longer-term storage, plate washers for processing plates, plate thermal sealers for applying heat seals, de-sealers for removing heat seals, or microplate incubators to ensure constant temperature during testing. Instrument companies have designed plate readers which can detect specific biological, chemical or physical events in samples stored in these plates. A specialized plate reader has also been developed which can perform quality control of microplate well contents, capable of identifying empty wells, filled wells and precipitate. The most common manufacturing process

3300-439: The technology is associated with the use of intact living organisms, like the nematode Caenorhabditis elegans and zebrafish ( Danio rerio ). In 2016-2018 plate manufacturers began producing specialized chemistry to allow for mass production of ultra-low adherent cell repellent surfaces which facilitated the rapid development of HTS amenable assays to address cancer drug discovery in 3D tissues such as organoids and spheroids;

3360-415: The wells of a stock plate to the corresponding wells of a completely empty plate. To prepare for an assay , the researcher fills each well of the plate with some biological entity that they wish to conduct the experiment upon, such as a protein , cells , or an animal embryo . After some incubation time has passed to allow the biological matter to absorb, bind to, or otherwise react (or fail to react) with

3420-541: The world can take advantage of this facility without lengthy intellectual property negotiations. With a compound library of over 200,000 small molecules, the MSSR has one of the largest compound deck of all universities on the west coast. Also, the MSSR features full functional genomics capabilities (genome wide siRNA, shRNA, cDNA and CRISPR) which are complementary to small molecule efforts: Functional genomics leverages HTS capabilities to execute genome wide screens which examine

3480-551: Was created in 1951 by a Hungarian, Dr. Gyula Takátsy , who machined six rows of 12 "wells" in Lucite . Subsequently, Dr. John Louis Sever modified the Hungarian design into a 96-well plate, which he published in 1962. However, common usage of the microplate began in the late 1980s when John Liner introduced a molded version. By 1990 there were more than 15 companies producing a wide range of microplates with different features. It

3540-545: Was estimated that 125 million microplates were used in 2000 alone. The word "Microtiter" is a registered trademark of Thermo Electron OY ( U.S. Trademark 754,087 .) Other trade names for microplates include Viewplate and Unifilter (introduced in the early 1990s by Polyfiltronics and sold by Packard Instrument, which is now part of PerkinElmer). In 1996, the Society for Biomolecular Screening (SBS), later known as Society for Biomolecular Sciences, began an initiative to create

3600-601: Was made by the Society for Biomolecular Sciences with the ANSI-Standards (ANSI/SBS 1-2004, ANSI/SBS 2-2004, ANSI/SBS 3-2004, ANSI/SBS 4-2004). These standards have been updated to and are now known as the ANSI SLAS standards. The ANSI SLAS microplate standards define a footprint, and a bottom flange geometry. These footprints & flanges are generally rigorously followed by all microplate manufacterers: Although

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