The field of vitamin D public health research has a pressing need to define sensitive and specific predictors of vitamin D status that can be used to determine whether an individual or population has a supply of vitamin D that is sufficient to meet requirements. The aim of this review is to highlight the considerations needed when evaluating evidence of the relations between vitamin D biomarkers and functional or health outcomes across the life cycle. It draws attention to the importance of distinguishing between biomarkers of supply, function, and outcome and of considering the many factors that could influence interpretation, such as life stage, ethnicity, body mass index, liver and kidney function, and dietary calcium and phosphorus intake. The vitamin D biomarkers that have shown the most utility to date are the plasma concentration of 25-hydroxyvitamin D (supply), the plasma concentration of parathyroid hormone (function), and the presence or absence of rickets (outcome). However, a single biomarker of vitamin D status or threshold value is unlikely to be valid in all situations. The field therefore needs research to refine existing biomarkers or establish new indicators that take the many factors into account and to identify useful functional biomarkers of vitamin D status for infants, children, women of reproductive age, and specific ethnic groups. However, evidence using the biomarkers currently available shows that frank vitamin D deficiency is a major public health problem in many parts of the world that requires urgent attention.