Since its initial report in 2009, the intestinal enteroid culture system has been a powerful tool used to study stem cell biology and development in the gastrointestinal tract. However, a major question is whether enteroids retain intestinal function and physiology. There have been significant contributions describing ion transport physiology of human intestinal organoid cultures, as well as physiology of gastric organoids, but critical studies on dietary fat absorption and chylomicron synthesis in primary intestinal enteroids have not been undertaken. Here we report that primary murine enteroid cultures recapitulate in vivo intestinal lipoprotein synthesis and secretion, and reflect key aspects of the physiology of intact intestine in regard to dietary fat absorption. We also show that enteroids can be used to elucidate intestinal mechanisms behind CVD risk factors, including tissue-specific apolipoprotein functions. Using enteroids, we show that intestinal apoC-III overexpression results in the secretion of smaller, less dense chylomicron particles along with reduced triacylglycerol secretion from the intestine.(jlr) This model significantly expands our ability to test how specific genes or genetic polymorphisms function in dietary fat absorption and the precise intestinal mechanisms that are critical in the etiology of metabolic disease.