## 4 Discussion  
Compared with conventional imaging techniques, IRT is advantageous as it can facilitate a quicker and more accurate diagnosis of physiological changes in ROIs [34]. Moreover, it is a noncontact, noninvasive diagnostic technique, which can measure physiological function by recording the exothermal status of the infrared rays emitted [35; 36; 37]. The skin temperature depends on the underlying tissue metabolism and local blood circulation. Therefore, the changes in physiological activity result in abnormal thermal patterns. Alterations in cutaneous blood flow and sweat patterns occur in the case of overload and inflammation [38]. Researchers suggest that ET obtained using IRT can be used to assess fever or stress in horses [15; 27; 39]. A higher ET may be correlated with ocular vasodilation, causing greater heat dissipation (identified using IRT), specifically in the lacimal caruncle, which contains capillaries controlled by the sympathetic nerves [15; 27; 39]. Although several studies have also shown that the medial canthus, particularly the lacimal caruncle, is an optimal location for measuring ET, there are no standards for measurement and analysis methods [21; 22; 23; 24]. [PERSON] et al. [22] obtained 791 images in 32 horses and measured the maximal ET in the medial posterior palpebral border of the lower eyelid and the lacimal caruncle in the circular area with a diameter of 1 cm. In addition, [PERSON] et al. [23] evaluated the maximum and mean eye ST at the commissure region of the lacimal caruncle in the circular area with a 22-pixels radius in 12 horses. However, recent efforts have attempted to designate a reference point or standardization method. [PERSON] et al. [21] determined whether the position of the IRT camera had an impact on recorded temperature and which camera position was optimal for indicating stress in 14 horses. This suggests that a 90\({}^{\circ}\) angle in relation to the sagittal plane in a mammal with anterolateral eyes is the optimal position for taking ET measurements using IRT. Further, [PERSON] et al. [33] studied the correlation between tympanic and facial temperatures in 508 humans. The results showed that the tympanic membrane temperature and medial canthus were the most similar. Moreover, a model was developed to predict tympanic temperature through statistical bias correction. These studies which attempted to define that standardization were incorporated into this study, resulting in higher accuracy and detail compared with previous studies that measured horse ET. In this study, the measurement area was localized to minimize unnecessary information, and the correlation between RT and ET measured at three areas in 172 horses (352 images) was quantitatively analyzed. The quantitative data obtained from IRT strongly influenced the ET result depending on the specific location of the measurement area; therefore, in this study, the 4 \(\times\) 4 pixel area, including the maximum temperature, was defined as the ROI. The maximum temperature was more correlated with RT compared with the average temperature of 4 \(\times\) 4 or 2 \(\times\) 2 pixels. However, the maximum temperature is a single pixel value and has a high efficacy in indicating the maximum or minimum value with a defective pixel. Therefore, this study suggests that the approach is only valid when the camera is properly calibrated and there is no image non-uniformity to prevent a single pixel value from not significantly affecting the outcome.