Faculty Opinions recommendation of High-throughput RNAi screening by time-lapse imaging of live human cells.

AbstractLive-imaging embryos in a high-throughput manner is essential for shedding light on a wide range of questions in developmental biology, but it is difficult and costly to mount and image embryos in consistent conditions. Here, we present OMMAwell, a simple, reusable device that makes it easy to mount up to hundreds of embryos in arrays of agarose microwells with customizable dimensions and spacing. OMMAwell can be configured to mount specimens for upright or inverted microscopes, and includes a reservoir to hold live-imaging medium to maintain constant moisture and osmolarity of specimens during time-lapse imaging. All device components can be cut from a sheet of acrylic using a laser cutter. Even a novice user will be able to cut the pieces and assemble the device in less than an hour. At the time of writing, the total materials cost is less than five US dollars. We include all device design files in a commonly used format, as well as complete instructions for its fabrication and use. We demonstrate a detailed workflow for designing a custom mold and employing it to simultaneously live-image dozens of embryos at a time for more than five days, using embryos of the cricket Gryllus bimaculatus as an example. Further, we include descriptions, schematics, and design files for molds that can be used with 14 additional vertebrate and invertebrate species, including most major traditional laboratory models and a number of emerging model systems. Molds have been user-tested for embryos including zebrafish (Danio rerio), fruit fly (Drosophila melanogaster), coqui frog (Eleutherodactylus coqui), annelid worm (Capitella teleta), amphipod crustacean (Parhyale hawaiensis), red flour beetle (Tribolium castaneum), and three-banded panther worm (Hofstenia miamia), as well mouse organoids (Mus musculus). Finally, we provide instructions for researchers to customize OMMAwell inserts for embryos or tissues not described herein.Summary StatementThis Techniques and Resources article describes an inexpensive, customizable device for mounting and live-imaging a wide range of tissues and species; complete design files and instructions for assembly are included.

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Low-Cost Automated Vectors and Modular Environmental Sensors for Plant Phenotyping

Sensors ◽

10.3390/s20113319 ◽

2020 ◽

Vol 20 (11) ◽

pp. 3319

Author(s):

Stuart A. Bagley ◽

Jonathan A. Atkinson ◽

Henry Hunt ◽

Michael H. Wilson ◽

Tony P. Pridmore ◽

...

Keyword(s):

High Throughput ◽

Low Cost ◽

Time Lapse ◽

Plant Phenotyping ◽

Environmental Sensors ◽

Controlled Conditions ◽

Sensor Platform ◽

High Throughput Phenotyping ◽

Controlled Environments ◽

Time Lapse Imaging

High-throughput plant phenotyping in controlled environments (growth chambers and glasshouses) is often delivered via large, expensive installations, leading to limited access and the increased relevance of “affordable phenotyping” solutions. We present two robot vectors for automated plant phenotyping under controlled conditions. Using 3D-printed components and readily-available hardware and electronic components, these designs are inexpensive, flexible and easily modified to multiple tasks. We present a design for a thermal imaging robot for high-precision time-lapse imaging of canopies and a Plate Imager for high-throughput phenotyping of roots and shoots of plants grown on media plates. Phenotyping in controlled conditions requires multi-position spatial and temporal monitoring of environmental conditions. We also present a low-cost sensor platform for environmental monitoring based on inexpensive sensors, microcontrollers and internet-of-things (IoT) protocols.

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Automated High-Throughput RNAi Screening in Human Cells Combined with Reporter mRNA Transfection to Identify Novel Regulators of Translation

PLoS ONE ◽

10.1371/journal.pone.0045943 ◽

2012 ◽

Vol 7 (9) ◽

pp. e45943 ◽

Cited By ~ 6

Author(s):

Claudia M. Casanova ◽

Peter Sehr ◽

Kerstin Putzker ◽

Matthias W. Hentze ◽

Beate Neumann ◽

...

Keyword(s):

High Throughput ◽

Human Cells ◽

Rnai Screening ◽

Mrna Transfection ◽

Reporter Mrna

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Time-Lapse Microscopy-Based Genome Wide RNAi Screening in Live Human Cells

3rd IEEE International Symposium on Biomedical Imaging: Macro to Nano, 2006. ◽

10.1109/isbi.2006.1624898 ◽

2006 ◽

Author(s):

B. Neumann ◽

M. Held ◽

U. Liebel ◽

H. Erfle ◽

P. Rogers ◽

...

Keyword(s):

Time Lapse ◽

Human Cells ◽

Rnai Screening ◽

Time Lapse Microscopy ◽

Genome Wide

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TIMING 2.0: high-throughput single-cell profiling of dynamic cell–cell interactions by time-lapse imaging microscopy in nanowell grids

Bioinformatics ◽

10.1093/bioinformatics/bty676 ◽

2018 ◽

Vol 35 (4) ◽

pp. 706-708 ◽

Cited By ~ 4

Author(s):

Hengyang Lu ◽

Jiabing Li ◽

Melisa A Martinez-Paniagua ◽

Irfan N Bandey ◽

Amit Amritkar ◽

...

Keyword(s):

User Interface ◽

High Throughput ◽

Graphical User Interface ◽

Time Lapse ◽

Previous Method ◽

Cell Interactions ◽

Supplementary Information ◽

Desktop Computer ◽

Time Lapse Imaging ◽

Cell Cell

Abstract Motivation Automated profiling of cell–cell interactions from high-throughput time-lapse imaging microscopy data of cells in nanowell grids (TIMING) has led to fundamental insights into cell–cell interactions in immunotherapy. This application note aims to enable widespread adoption of TIMING by (i) enabling the computations to occur on a desktop computer with a graphical processing unit instead of a server; (ii) enabling image acquisition and analysis to occur in the laboratory avoiding network data transfers to/from a server and (iii) providing a comprehensive graphical user interface. Results On a desktop computer, TIMING 2.0 takes 5 s/block/image frame, four times faster than our previous method on the same computer, and twice as fast as our previous method (TIMING) running on a Dell PowerEdge server. The cell segmentation accuracy (f-number = 0.993) is superior to our previous method (f-number = 0.821). A graphical user interface provides the ability to inspect the video analysis results, make corrective edits efficiently (one-click editing of an entire nanowell video sequence in 5–10 s) and display a summary of the cell killing efficacy measurements. Availability and implementation Open source Python software (GPL v3 license), instruction manual, sample data and sample results are included with the Supplement (https://github.com/RoysamLab/TIMING2). Supplementary information Supplementary data are available at Bioinformatics online.

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Cleavage of Human Embryos: Options and Diversity

Acta Naturae ◽

10.32607/20758251-2016-8-3-88-96 ◽

2016 ◽

Vol 8 (3) ◽

pp. 88-96

Author(s):

Yu. K. Doronin ◽

I. V. Senechkin ◽

L. V. Hilkevich ◽

M. A. Kurcer

Keyword(s):

Time Lapse ◽

Reproductive Technologies ◽

Human Embryos ◽

Imaging Data ◽

Noninvasive Methods ◽

Topographic Features ◽

Time Parameters ◽

Embryo Cleavage ◽

Time Lapse Imaging ◽

Developmental Dynamics

In order to estimate the diversity of embryo cleavage relatives to embryo progress (blastocyst formation), time-lapse imaging data of preimplantation human embryo development were used. This retrospective study is focused on the topographic features and time parameters of the cleavages, with particular emphasis on the lengths of cleavage cycles and the genealogy of blastomeres in 2- to 8-cell human embryos. We have found that all 4-cell human embryos have four developmental variants that are based on the sequence of appearance and orientation of cleavage planes during embryo cleavage from 2 to 4 blastomeres. Each variant of cleavage shows a strong correlation with further developmental dynamics of the embryos (different cleavage cycle characteristics as well as lengths of blastomere cycles). An analysis of the sequence of human blastomere divisions allowed us to postulate that the effects of zygotic determinants are eliminated as a result of cleavage, and that, thereafter, blastomeres acquire the ability of own syntheses, regulation, polarization, formation of functional contacts, and, finally, of specific differentiation. This data on the early development of human embryos obtained using noninvasive methods complements and extend our understanding of the embryogenesis of eutherian mammals and may be applied in the practice of reproductive technologies.

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