A Pillar-Based High-Throughput Myogenic Differentiation Assay to Assess Drug Safety

High-throughput, pillar-strip-based assays have been proposed as a drug-safety screening tool for developmental toxicity. In the assay described here, muscle cell culture and differentiation were allowed to occur at the end of a pillar strip (eight pillars) compatible with commercially available 96-well plates. Previous approaches to characterize cellular differentiation with immunostaining required a burdensome number of washing steps; these multiple washes also resulted in a high proportion of cellular loss resulting in poor yield. To overcome these limitations, the approach described here utilizes cell growth by easily moving the pillars for washing and immunostaining without significant loss of cells. Thus, the present pillar-strip approach is deemed suitable for monitoring high-throughput myogenic differentiation. Using this experimental high-throughput approach, eight drugs (including two well-known myogenic inhibitory drugs) were tested at six doses in triplicate, which allows for the generation of dose–response curves of nuclei and myotubes in a 96-well platform. As a result of comparing these F-actin (an actin-cytoskeleton protein), nucleus, and myotube data, two proposed differentiation indices—curve-area-based differentiation index (CA-DI) and maximum-point-based differentiation index (MP-DI) were generated. Both indices successfully allowed for screening of high-myogenic inhibitory drugs, and the maximum-point-based differentiation index (MP-DI) experimentally demonstrated sensitivity for quantifying drugs that inhibited myogenic differentiation.

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High-Throughput Affinity Selection Mass Spectrometry Using SAMDI-MS to Identify Small-Molecule Binders of the Human Rhinovirus 3C Protease

SLAS DISCOVERY Advancing Life Sciences ◽

10.1177/24725552211023211 ◽

2021 ◽

pp. 247255522110232

Author(s):

Michael D. Scholle ◽

Doug McLaughlin ◽

Zachary A. Gurard-Levin

Keyword(s):

Mass Spectrometry ◽

Drug Discovery ◽

High Throughput ◽

High Throughput Screening ◽

Human Rhinovirus ◽

Binding Potential ◽

Affinity Selection ◽

Response Curves ◽

Desorption Ionization ◽

Self Assembled

Affinity selection mass spectrometry (ASMS) has emerged as a powerful high-throughput screening tool used in drug discovery to identify novel ligands against therapeutic targets. This report describes the first high-throughput screen using a novel self-assembled monolayer desorption ionization (SAMDI)–ASMS methodology to reveal ligands for the human rhinovirus 3C (HRV3C) protease. The approach combines self-assembled monolayers of alkanethiolates on gold with matrix-assisted laser desorption ionization time-of-flight (MALDI TOF) mass spectrometry (MS), a technique termed SAMDI-ASMS. The primary screen of more than 100,000 compounds in pools of 8 compounds per well was completed in less than 8 h, and informs on the binding potential and selectivity of each compound. Initial hits were confirmed in follow-up SAMDI-ASMS experiments in single-concentration and dose–response curves. The ligands identified by SAMDI-ASMS were further validated using differential scanning fluorimetry (DSF) and in functional protease assays against HRV3C and the related SARS-CoV-2 3CLpro enzyme. SAMDI-ASMS offers key benefits for drug discovery over traditional ASMS approaches, including the high-throughput workflow and readout, minimizing compound misbehavior by using smaller compound pools, and up to a 50-fold reduction in reagent consumption. The flexibility of this novel technology opens avenues for high-throughput ASMS assays of any target, thereby accelerating drug discovery for diverse diseases.

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Automatic 3D Cell Analysis in High-Throughput Microarray Using Micropillar and Microwell Chips

CrossRef Listing of Deleted DOIs ◽

10.1177/1087057115597635 ◽

2015 ◽

Vol 20 (9) ◽

pp. 1178-1184 ◽

Cited By ~ 17

Author(s):

Dong Woo Lee ◽

Moo-Yeal Lee ◽

Bosung Ku ◽

Do-Hyun Nam

Keyword(s):

Standard Deviation ◽

High Throughput ◽

High Throughput Screening ◽

Doubling Time ◽

Cultured Cells ◽

Measurement Methods ◽

Cell Analysis ◽

Response Curves ◽

U251 Cell ◽

Doubling Times

Area-based and intensity-based 3D cell viability measurement methods are compared in high-throughput screening in order to analyze their effects on the assay results (doubling time and IC50) and their repeatability. Many other 3D cell-based high-throughput screening platforms had been previously introduced, but these had not clearly addressed the effects of the two methods on the assay results and assay repeatability. In this study, the optimal way to analyze 3D cultured cells is achieved by comparing day-to-day data of doubling times and IC50 values obtained from the two methods. In experiments, the U251 cell line is grown in chips. The doubling time, based on the area of the 3D cells, was 27.8 ± 1.8 h (standard deviation: 6.6%) and 27.8 ± 3.8 h (standard deviation: 13.7%) based on the intensity of the 3D cells. The doubling time calculated by area shows a smaller standard deviation than one calculated by intensity. IC50 values calculated by both methods are very similar. The standard deviations of IC50 values for the two methods were within ±3-fold. The IC50 variations of the 12 compounds were similar regardless of the viability measurement methods and were highly related to the shape of the dose–response curves.

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Development of a high-throughput thermoelectric screening tool for combinatorial thin film libraries

Applied Surface Science ◽

10.1016/j.apsusc.2007.05.091 ◽

2007 ◽

Vol 254 (3) ◽

pp. 765-767 ◽

Cited By ~ 31

Author(s):

M. Otani ◽

K. Itaka ◽

W. Wong-Ng ◽

P.K. Schenck ◽

H. Koinuma

Keyword(s):

Thin Film ◽

High Throughput ◽

Screening Tool

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A paradigm approach toward predicting barotrauma

Indian Journal of Aerospace Medicine ◽

10.25259/ijasm_2019_9 ◽

2019 ◽

Vol 63 ◽

pp. 44-48

Author(s):

V Raghunandan ◽

SS Mohapatra

Keyword(s):

Eustachian Tube ◽

Middle Ear ◽

Screening Tool ◽

Significant Loss ◽

Causal Factor ◽

Step Test ◽

Medical Problems ◽

And Training ◽

Nose And Throat

Barotrauma is one of the most common medical problems associated with aviation and has been a causal factor for significant loss of trained aircrew hours. The occurrence of barotrauma has been reported not only in flight but also during simulated exposures to changing pressures as a part of aeromedical evaluation and training. This occurrence is despite the screening of individuals by otoscopy and tympanometry before exposure to barometric changes. In such cases, baro-challenge-induced eustachian tube (ET) dysfunction has been identified as the most probable cause, leading to decreased ability to equalize pressures across the middle ear. This paper describes a paradigm approach to the prediction of baro-challenge-induced ET dysfunction using the nine-step test and ET dysfunction questionnaire. The inclusion of the test as a screening tool before exposure of aircrew to baro-challenge after recovery from their ear, nose, and throat ailments is recommended to prevent occurrence/ reoccurrence of barotrauma.

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Detection of N-acyl homoserine lactones using a traI-luxCDABE-based biosensor as a high-throughput screening tool

BMC Biotechnology ◽

10.1186/1472-6750-8-59 ◽

2008 ◽

Vol 8 (1) ◽

pp. 59 ◽

Cited By ~ 13

Author(s):

Steve P Bernier ◽

Anne L Beeston ◽

Pamela A Sokol

Keyword(s):

High Throughput ◽

High Throughput Screening ◽

Screening Tool ◽

Homoserine Lactones ◽

Acyl Homoserine Lactones

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Chemically diverse polymer microarrays and high throughput surface characterisation: a method for discovery of materials for stem cell culture

Biomaterials Science ◽

10.1039/c4bm00054d ◽

2014 ◽

Vol 2 (11) ◽

pp. 1604-1611 ◽

Cited By ~ 27

Author(s):

A. D. Celiz ◽

J. G. W. Smith ◽

A. K. Patel ◽

R. Langer ◽

D. G. Anderson ◽

...

Keyword(s):

Cell Culture ◽

Stem Cell ◽

High Throughput ◽

Screening Tool ◽

New Materials ◽

Surface Characterisation ◽

Stem Cell Culture

Chemically diverse polymer microarrays as a powerful screening tool for the discovery of new materials for a variety of applications.

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Predictive Models of Prenatal Developmental Toxicity from ToxCast High-Throughput Screening Data

Toxicological Sciences ◽

10.1093/toxsci/kfr220 ◽

2011 ◽

Vol 124 (1) ◽

pp. 109-127 ◽

Cited By ~ 140

Author(s):

Nisha S. Sipes ◽

Matthew T. Martin ◽

David M. Reif ◽

Nicole C. Kleinstreuer ◽

Richard S. Judson ◽

...

Keyword(s):

High Throughput ◽

Predictive Models ◽

High Throughput Screening ◽

Developmental Toxicity

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Copper Oxide Nanoparticle-Based Immunosensor for Zearalenone Analysis by Combining Automated Sample Pre-Processing and High-Throughput Terminal Detection

Sensors ◽

10.3390/s21196538 ◽

2021 ◽

Vol 21 (19) ◽

pp. 6538

Author(s):

Zhihong Xuan ◽

Yanxiang Wu ◽

Hongmei Liu ◽

Li Li ◽

Jin Ye ◽

...

Keyword(s):

High Throughput ◽

Screening Tool ◽

Detection Method ◽

Sample Pretreatment ◽

Fluorescence Signal ◽

Signal Conversion ◽

Cuo Nps ◽

Acidic Conditions ◽

Oxide Nanoparticle ◽

Efficient Screening

A rapid and high-throughput fluorescence detection method for zearalenone (ZEN) based on a CuO nanoparticle (NP)-assisted signal amplification immunosensor was developed using an automated sample pretreatment and signal conversion system. CuO NPs with high stability and biocompatibility were used as carriers to immobilize anti-ZEN antibodies. The obtained CuO NP-anti-ZEN can maintain the ability to recognize target toxins and act as both a signal source and carrier to achieve signal conversion using automated equipment. In this process, target toxin detection is indirectly transformed to Cu2+ detection because of the large number of Cu2+ ions released from CuO NPs under acidic conditions. Finally, a simple and high-throughput fluorescence assay based on a fluorescent tripeptide molecule was employed to detect Cu2+, using a multifunctional microporous plate detector. A good linear relationship was observed between the fluorescence signal and the logarithm of ZEN concentration in the range of 16.0–1600.0 μg/kg. Additionally, excellent accuracy with a high recovery yield of 99.2–104.9% was obtained, which was concordant with the results obtained from LC-MS/MS of naturally contaminated samples. The CuO NP-based assay is a powerful and efficient screening tool for ZEN detection and can easily be modified to detect other mycotoxins.

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