Optimization of Experimental Settings for the Assessment of Reactive Oxygen Species Production by Human Blood


Journal article


Tânia R P Soares, Daniela Rodrigues, M. Sarraguça, Sílvia Rocha, J. Lima, Daniela Ribeiro, E. Fernandes, M. Freitas
Oxidative medicine and cellular longevity, 2019

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APA   Click to copy
Soares, T. R. P., Rodrigues, D., Sarraguça, M., Rocha, S., Lima, J., Ribeiro, D., … Freitas, M. (2019). Optimization of Experimental Settings for the Assessment of Reactive Oxygen Species Production by Human Blood. Oxidative Medicine and Cellular Longevity.


Chicago/Turabian   Click to copy
Soares, Tânia R P, Daniela Rodrigues, M. Sarraguça, Sílvia Rocha, J. Lima, Daniela Ribeiro, E. Fernandes, and M. Freitas. “Optimization of Experimental Settings for the Assessment of Reactive Oxygen Species Production by Human Blood.” Oxidative medicine and cellular longevity (2019).


MLA   Click to copy
Soares, Tânia R. P., et al. “Optimization of Experimental Settings for the Assessment of Reactive Oxygen Species Production by Human Blood.” Oxidative Medicine and Cellular Longevity, 2019.


BibTeX   Click to copy

@article{t2019a,
  title = {Optimization of Experimental Settings for the Assessment of Reactive Oxygen Species Production by Human Blood},
  year = {2019},
  journal = {Oxidative medicine and cellular longevity},
  author = {Soares, Tânia R P and Rodrigues, Daniela and Sarraguça, M. and Rocha, Sílvia and Lima, J. and Ribeiro, Daniela and Fernandes, E. and Freitas, M.}
}

Abstract

The purpose of an experimental design is to improve the productivity of experimentation. It is an efficient procedure for planning experiments, so the data obtained can be analyzed to yield a valid and objective conclusion. This approach has been used as an important tool in the optimization of different analytical approaches. A D-optimal experimental design was used here, for the first time, to optimize the experimental conditions for the detection of reactive oxygen species (ROS) produced by human blood from healthy donors, a biological matrix that better resembles the physiologic environment, following stimulation by a potent inflammatory mediator, phorbol-12-myristate-13-acetate (PMA). For that purpose, different fluorescent probes were used, as 2′,7′-dichlorodihydrofluorescein diacetate (DCFH-DA), 2-[6-(4′-amino)-phenoxy-3H-xanthen-3-on-9-yl] benzoic acid (APF), and 10-acetyl-3,7-dihydroxyphenoxazine (amplex red). The variables tested were the human blood dilution, and the fluorescent probe and PMA concentrations. The experiments were evaluated using the Response Surface Methodology and the method was validated using specific compounds. This model allowed the search for optimal conditions for a set of responses simultaneously, enabling, from a small number of experiments, the evaluation of the interaction between the variables under study. Moreover, a cellular model was implemented and optimized to detect the production of ROS using a yet nonexplored matrix, which is human blood.


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