Stress Concepts and Applications in Various Matrices with a Focus on Hair Cortisol and Analytical Methods

Simple Summary Stress in domestic animals can lead to serious consequences. We reviewed biological specimens (fluid and non-fluid types), that are capable of evaluating either cortisol or corticosterone steroid levels as major biomarkers of stress (acute or chronic) in different animal species. Proper methods to evaluate the chronic stress of animals through hormonal analysis can save time and cost of experiments in clinical laboratories due to different research groups applying different techniques for preparation, extraction, and analysis of the biological specimens that may result in fluctuating and inaccurate values. In addition, we reviewed the immunoassays such as singleplex immunoassays (ELISA) and multiplex immunoassays as the most common method used for detecting multi-biomarkers of stress. In this review, we aimed to: (1) explain the classification of stress, (2) discuss the matrices that can be used as biomarkers of stress, their comparison, and limitations, and present the most important reliable matrix, (3) compare the analytical methods for measuring stress hormones after sample preparation to determine the result. In this review, we have discussed the method of sample collection, sex and age effects, the body regions to be selected, and the method of analysis of cortisol and corticosterone in different body indices that may be used from time to time. We also compared the advantages and disadvantages of each matrix and technique for analyzing stress hormones. When studying stress in animals, it is important to understand the types of stress and their classification, and how to assess the stress levels in different animal species using different matrices accurately and precisely. The classification of stress types helps to distinguish between good stress (eustress) and bad stress (distress). Hence, first, it is crucial to assess the animal's level of stress in a non-intrusive manner and second to identify the type of stress that is best suited to its environment. Third, it is also important to analyze the obtained samples using a suitable method to increase the validity of stress hormone measurements. Therefore, in this review, we aim to: (1) explain the classification of stress, (2) discuss the wide range of body matrices (e.g., saliva, milk, hair, urine, feces, sweat, fins, etc.) that can be used as samples to evaluate stress levels, as well as their comparisons and limitations, and present the reliable matrices for measuring stress hormones with special emphasis on hair, (3) compare the analytical methods for measuring stress hormones after sample preparation. Despite some literature that does not include hair as a reliable matrix for evaluating stress levels, hair is one of the matrices for measuring long-term stress hormone accumulations. This review discusses some factors that influence the level of stress hormones in the hair. By understanding these issues, the scientific community will not only be able to improve the understanding of stress and biomarker evaluation but also suggest how to deal with the consequences of stress in future research.