A series of studies were conducted to ascertain the importance of autotrophic picoplankton (0.2 - 2.0 fm in the longest dimension) in freshwaters. These included surveys of populations in lakes and rivers, and experiments on strains which had been isolated in clonal, axenic culture. Methods were developed to preserve, count and identify autotrophic picoplankton. Epifluorescence microscopy was used routinely throughout the study and it was shown that cells could be preserved in buffered formalin or glutaraldehyde for at least 12 months without a loss in cell count. Autotrophic picoplankton fluoresced red or orange depending on the photosynthetic pigments present, and a method based on fluorescence characteristics was developed to distinguish cyanobacterial from eukaryotic cells. Eukaryotes fluoresced most intensely under blue excitation light and were barely visible under green excitation light and cyanobacterial cells had the reverse response. In addition, fluorescence from eukaryotic cells faded faster than the fluorescence from cyanobacterial cells. Samples from standing waters from around the world were collected from 5 continents in a survey to discover how widespread autotrophic picoplankton are in the world; densities ranged from 1.02 x 10(^6) from a pool in Saudi Arabia to 1.20 x 10(^6) in Ennerdale Water in the UK. A survey of 30 lakes in northern England and Scotland revealed autotrophic picoplankton in every sample. Cell densities ranged from a minimum of 1.02 x 10(^2) in Esthwaite Water to a maximum of 4.26 x 10(^5) in Ennerdale Water. In two mountain streams the only autotrophic picoplankton found were aberrant. Studies on the seasonality of autotrophic picoplankton from 10 UK lakes showed that population densities differed by at least two orders of magnitude in a year. In every lake densities reached a maximum at mid- to late summer and highest densities were found in nutrient-poor waters. There was a marked difference in the relative abundance of orange and red fluorescing cells in all lakes. A number of standard methods were used to isolate and purify strains of autotrophic picoplankton, including a novel method using a laser flow cytometer to purify bacterized unialgal cultures. 14 strains were studied (9 obtained the author) and they demonstrated different growth rates when grown with different nitrogen substrates and when grown under different photon flux density. Twelve strains showed cell-bound phosphomonoesterase activity at pH 10.3 and nine strains showed cell bound phosphodiesterase activity at pH 10.3. No strain showed evidence of nitrogen fixation, chemoheterotrophic growth, desiccation tolerance, motility nor chromatic adaptation.