TY - JOUR
T1 - Technique for Determining the Viability of Acanthamoeba Cysts Treated with a Cysticidal Agent Based on Membrane Integrity
AU - Che Mohamad, Nor Farah Azwani
AU - Failei, Nur Syakinah Nafisa
AU - Mohd Rased, Nurhidayana
AU - Adnan, Azila
AU - Ling, Ma Nyuk
AU - Zakeri, Hazlina Ahamad
AU - Kusrini, Eny
AU - Hashim, Fatimah
N1 - Funding Information:
This study was funded by UMT TAPE-RG 55115 (Talent and Publication Enhancement Research Grant). The facilities were supported by the Institute of Marine Biotechnology, Universiti Malaysia Terengganu.
Publisher Copyright:
© 2021
PY - 2021/10
Y1 - 2021/10
N2 - 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.
AB - 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.
KW - Dithiothreitol
KW - Fluorescence intensity
KW - High content screening
KW - Keratitis
KW - Propidium iodide
UR - http://www.scopus.com/inward/record.url?scp=85117884011&partnerID=8YFLogxK
U2 - 10.14716/ijtech.v12i4.4207
DO - 10.14716/ijtech.v12i4.4207
M3 - Article
AN - SCOPUS:85117884011
SN - 2086-9614
VL - 12
SP - 720
EP - 727
JO - International Journal of Technology
JF - International Journal of Technology
IS - 4
ER -