Multiwavelength ultraviolet–visible (UV–Vis) transmission spectroscopy is a relatively simple technique that can provide considerable quantitative information on the properties of micron and submicron particle suspensions. Two important particle properties are particle size distribution (PSD) and chemical composition. These properties provide characteristics for the identification and classification of biological systems ranging in size and composition from proteins and nucleic acids to cells. By measuring the complete UV–Vis spectrum, the combined scattering and absorption properties are obtained as a function of wavelength. The quantitative evaluation of the size distribution and chemical composition is accomplished through the application of light-scattering theory. This paper reports on the estimation of the optical properties of human blood platelets and their use in the interpretation of platelet UV–Vis spectra within the context of Mie theory. The model developed herein provides reliable and accurate estimates for the PSD and particle number of platelet suspensions. One potential application of this characterization method is in the analysis of platelet activation by thrombin. Quantification of spectral data with respect to average particle size and particle number provides a real-time description of the dramatic changes that accompany the platelet activation process.