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PLoS One. 2018 Oct 5;13(10):e0203597. doi: 10.1371/journal.pone.0203597. eCollection 2018.

Design and validation of an open-source modular Microplate Photoirradiation System for high-throughput photobiology experiments.

Author information

1
Department of Cell, Developmental and Regenerative Biology, Black Family Stem Cell Institute, Icahn School of Medicine at Mount Sinai, New York, NY, 10029 United States of America.
2
Department of Neurosurgery, Sinai Biodesign, Icahn School of Medicine at Mount Sinai, New York, New York, 10029 United States of America.
3
Merck Engineering LLC, Hamburg, NJ, 07495 United States of America.
4
Department of Pediatrics, Allergy and Immunology, Icahn School of Medicine at Mount Sinai, New York, NY, USA, Icahn School of Medicine at Mount Sinai, New York, NY, 10029 United States of America.
5
Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA, Icahn School of Medicine at Mount Sinai, New York, NY, 10029 United States of America.
6
Department of Pharmacological Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, United States of America.
7
Biofisika Institute (CSIC, UPV/EHU), University of the Basque Country, Leioa, Spain.

Abstract

Research in photobiology is currently limited by a lack of devices capable of delivering precise and tunable irradiation to cells in a high-throughput format. This limits researchers to using expensive commercially available or custom-built light sources which make it difficult to replicate, standardize, optimize, and scale experiments. Here we present an open-source Microplate Photoirradiation System (MPS) developed to enable high-throughput light experiments in standard 96 and 24-well microplates for a variety of applications in photobiology research. This open-source system features 96 independently controlled LEDs (4 LEDs per well in 24-well), Wi-Fi connected control and programmable graphical user interface (GUI) for control and programming, automated calibration GUI, and modular control and LED boards for maximum flexibility. A web-based GUI generates light program files containing irradiation parameters for groups of LEDs. These parameters are then uploaded wirelessly, stored and used on the MPS to run photoirradiation experiments inside any incubator. A rapid and semi-quantitative porphyrin metabolism assay was also developed to validate the system in wild-type fibroblasts. Protoporphyrin IX (PpIX) fluorescence accumulation was induced by incubation with 5-aminolevulinic acid (ALA), a photosensitization method leveraged clinically to destroy malignant cell types in a process termed photodynamic therapy (PDT), and cells were irradiated with 405nm light with varying irradiance, duration and pulsation parameters. Immediately after light treatment with the MPS, subsequent photobleaching was measured in live, adherent cells in both 96-well and a 24-well microplates using a microplate reader. Results demonstrate the utility and reliability of the Microplate Photoirradiation System to irradiate cells with precise irradiance and timing parameters in order to measure PpIx photobleaching kinetics in live adherent cells and perform comparable experiments with both 24 and 96 well microplate formats. The high-throughput capability of the MPS enabled measurement of enough irradiance conditions in a single microplate to fit PpIX fluorescence to a bioexponential decay model of photobleaching, as well as reveal a dependency of photobleaching on duty-cycle-but not frequency-in a pulsed irradiance regimen.

PMID:
30289930
PMCID:
PMC6173374
DOI:
10.1371/journal.pone.0203597
[Indexed for MEDLINE]
Free PMC Article

Conflict of interest statement

We have the following interests: Chris Merck is affiliated with his own LLC (Merck Engineering LLC). Merck Engineering LLC did not contribute funding to the development of the MPS or its biological validation and has no pecuniary interest in this study. Chris Merck worked as a volunteer collaborator not representing any company or institution. There are no patents, products in development or marketed products to declare. This does not alter our adherence to all the PLOS ONE policies on sharing data and materials.

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