Fluorescence microscopy has played an essential role in cell biology and immunology by allowing visualization of specific molecular components of cells. Recently, confocal laser scanning microscopy and atomic force microscopy have been introduced into the field and have extraordinarily changed the strategy for studying the dynamic molecular organization and cell signaling. They will bring about a new age in cell biology and immunology. Thus, we will review our recent work for cell signaling and imaging in mast cell activation, intending for many researchers to obtain valuable information to study the cell function in allergy and clinical immunology.