Under optimal conditions, plasma cholesterol homeostasis is maintained by a variety of mechanisms, balancing input and output, thereby preventing the net accumulation of cholesterol in circulation and tissues. Among these mechanisms, intestinal cholesterol absorption has recently re-emerged as a potentially important contributor to cholesterol homeostasis. However, its regulation has been difficult to study in humans because of technical limitations in methodologies. In this review the major methods available for measuring cholesterol absorption including those that utilize cholesterol balance, single dose isotopic feeding, dual isotope plasma ratio, continuous isotope feeding, intestinal perfusion, stable isotopes and serum plant sterols or cholestanol to cholesterol ratios are reviewed and contrasted. Emphasis is placed on the strengths, technical and interpretational limitations and their applicability for use in metabolic, small-scale outpatient, population and large-scale intervention studies.