Stable isotope and ecological stoichiometry investigations of aquatic food webs require separate measurements of microalgae and detritus, but fine particulate matter collected as seston or scraped from biofilms typically is an unknown mixture of these two components plus other material. This paper describes an economical method to partition fine particulate matter into predominantly algal and detrital components by centrifugation in colloidal silica. Centrifugation using this density also worked well to separate algae from inorganic matter. The protocol was tested by sampling a broad range of water bodies in Queensland, Australia. Seston was concentrated using an inexpensive continuous centrifuge and organic matter was collected from surfaces of rocks, mud, or plants. Separation in colloidal silica was achieved using a standard benchtop centrifuge. When the colloidal silica was adjusted to a density of 1.27 g/cm3, the light fraction (supernatant) tended to be dominated by algal organic matter while the heavy fraction tended to be more detrital. Ratios of organic carbon to chlorophyll a indicated the efficacy of the separations. Stable C and N isotope ratios often differed considerably between the algal and detrital fractions, demonstrating the need to perform such separations in many aquatic environments.