This paper provides a novel technique to detect transparent biological living cells trapped in a microfluidic MEMS device by optical diffraction. The device essentially consists of an optical aperture or an aperture array patterned in metal layer and a microfluidic chamber positioned above the center of the aperture. When the cells in the chamber are illuminated through the aperture, the far-field diffraction pattern can be recorded by a CCD camera or a photodetector array. This diffraction pattern uniquely corresponds to the sizes, positions, and intrinsic optical properties of the aperture, cells, and the microfluidic chamber materials, so any unknown but relevant parameter is able to be extrapolated when all other parameters are fixed or identified. This paper describes in detail the designs of various microfluidic chambers and apertures for this application, and the development of a complete set of software for the analysis of the cells' optical properties. Compared with other currently available methods for the detection of transparent living cells, this method has the advantages of simple device structure, easy to manipulate, able to simultaneously detect several cells of different species, as well as providing accurate and sensitive results. Besides the detection of living cells, this technique can also be used to detect or characterize other transparent or low optical absorption particles, such as polymer spheres or insoluble droplets, inside an aqueous solution.