Technique for Determining the Viability of Acanthamoeba Cysts Treated with a Cysticidal Agent Based on Membrane Integrity
This study presents a straightforward and reliable method for determining the viability of Acanthamoeba cysts. A standard method for determining Acanthamoeba cyst viability in an in vitro cytotoxicity analysis is required to ensure that the double-walled and sturdy cysts are affected by the sub...
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| Autores principales: | , , , , , , , |
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| Formato: | article |
| Lenguaje: | EN |
| Publicado: |
Universitas Indonesia
2021
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| Materias: | |
| Acceso en línea: | https://doaj.org/article/d03f68f8d3e045cab3bd4686ed4a5c67 |
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| Sumario: | This study presents a
straightforward and reliable method for determining the viability of Acanthamoeba
cysts. A standard method for determining Acanthamoeba cyst viability in
an in vitro cytotoxicity analysis is
required to ensure that the double-walled and sturdy cysts are affected by the
substance tested. In this study, a new approach was used to determine the
cysticidal potential of redox Cleland’s reagent, dithiothreitol (DTT), against Acanthamoeba
cysts. This approach constitutes a significant breakthrough, as the cyst form
of Acanthamoeba is known for its high resistance to various chemicals
and drugs used to treat infections of the central nervous system and eyes
caused by Acanthamoeba. Cyst viability was evaluated based on the
intensity of the cyst population under fluorescence produced by propidium
iodide (PI) dye and measured using an enzyme-linked immunosorbent assay (ELISA)
reader at an absorbance of 636 nm. The results were validated using
high-content screening (HCS). For analysis, an individual cell was imaged and
examined for phenotypic changes in the Acanthamoeba cyst at the cyst
population level. Fluorescence intensity of the cysts in each well in a 96-well
plate was measured using Image J software. HCS is an automated technique that
uses fluorescence microscopy to produce quantitative data. |
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