Mast cells have long been known to play a detrimental role in the pathogenesis of IgE-associated allergic disorders by their ability to release a wide variety of pro-inflammatory mediators. A number of studies, however, have demonstrated that mast cells play a beneficial role in innate host defense against bacterial infections. Since mast cells clearly play both physiological and pathophysiological functions in the body, it is important to learn about the components of mast cells that drive these responses. The functional roles of mast cell in vivo have been principally characterized by comparing the biological responses in mast cell-deficient mice (WBB6F(1)-W/W(v)), their normal wild-type littermates (WBB6F(1)-+/+) and mast cell deficient mice reconstituted locally or systemically with mast cells cultured from the bone marrow cells of WBB6F(1)-+/+ mice (WBB6F(1)-W/W(v)+MC). Recently investigators have demonstrated that mast cell-deficient mice (WBB6F(1)-W/W(v)) can be reconstituted with mast cells derived in vitro from the bone marrow cells of certain gene knock-out mice or genetically-manipulated embryonic stem cells. This novel approach of analyzing the biological consequences of gene mutations in mast cells will help us to better understand the role of individual gene products in mast cell responses. In this review, we discuss these new approaches to investigate the functions of mast cells in vivo.