Steroid hormones have been proposed to influence the development of lateralization of the brain and behavior. We briefly describe the available hypotheses explaining this influence. These are all based on human data, whereas experimental testing is almost exclusively limited to other animal models. As a consequence, different research fields use different techniques and study species when to investigate the relationship between steroid hormones and lateralization. The aim of this chapter is to present an overview of available techniques to study this relationship with an interdisciplinary approach. To this end, we describe the basics of hormone secretion and mechanisms of action for androgens, estrogens, progesterone, and corticosteroids. Next, general issues related to hormone sampling and hormone assays are discussed. We then present a critical overview of correlational and experimental methods to study the influence of prenatal and postnatal hormones on lateralization. These methods include hormone measurement in amniotic fluid, saliva, urine, feces, and blood plasma or serum of the fetus, mother, and umbilical cord. We also discuss hormone-mediated maternal effects, the manipulation of hormone levels in the embryo or mother, hormone treatment in persons with Gender Dysphoria, and the 2D:4D finger length ratio as a proxy for prenatal testosterone exposure. Additionally, we discuss the prerequisites for hormones to act in the brain and induce behavioral lateralization, a topic that has not yet received sufficient attention. We argue that lateralization can and should be studied at different levels of organization, namely, structural and functional brain lateralization, perception and cognition, lateralized motor output, and performance. We present tests for these different levels and argue that keeping these levels apart is important as well as realizing that lateralization and the hormonal influence on it may be different at different levels for different functions and different species. We conclude that the study of hormonal influences on lateralization of brain and behavior has not yet exploited the knowledge and wide array of techniques currently available, leaving an interesting research field substantially under-explored.