Salivette synthetic swabs

Hydrofluoric acid (HF) was purchased from Technic Inc. (Saint-Denis, France). Nitric acid (HNO₃) came from Lach-Ner Ltd. (Neratovice, Czech Republic). Silver nitrate (AgNO₃), reduced L-glutathione, 2-thiobarbituric acid, and malondialdehyde tetrabutylammonium salt were purchased from Sigma-Aldrich (Taufkirchen, Germany). Crystal violet (CV) was obtained from Acros Organics (Geel, Belgium). Salivette^® synthetic swabs came from Sarstedt (Numbrecht, Germany). The water used throughout the study was distilled.

To measure stress during the stress paradigm, salivary cortisol will be collected at six time points (see Fig. 4). Salivette synthetic swabs (Sarstedt, Germany), specifically designed for salivary cortisol analysis, will be used. The swabs will be centrifuged for 4 min at 3000 rpm (Jouan CR412 centrifuge), and the filtrates will be stored at − 80 °C [100 (link), 143 ]. The routine laboratory of the Ghent University Hospital will assay the salivary cortisol by means of a Modular E 170 immunoanalyzer system (Roche Diagnostics, Mannheim, Germany) using the Roche Cobas Cortisol assay. For a more detailed description of this analysis technique, see van Aken, Romijn, Miltenburg, and Lentjes [139 (link)]. Next, the cortisol concentrations from the analysis (μg/dL) will be multiplied with a conversion factor of 27.586, resulting in the SI units nanomoles per liter (nmol/L) [41 (link)]. The difference in the cortisol level throughout the five blocks (stress paradigm) will represent the adaptation or maladaptation to the stress and thus act as a proxy of eustress and distress (see Fig. 1).