An Particle size distribution (PSD) is a fundamental physical property, which should be measured to understand the role of the colloids in the biogeochemical cycle of aquatic contaminants. The single particle counter (SPC) is an instrument designed to precisely determine sub-micron PSD. The advantages of this system are: to give an absolute colloid number concentration, to measure without bias highly polydispersed samples, to avoid artefacts resulting from sample preconcentration, to be transportable, allowing on-site measurements. This study presents the results of the methodological work done on our SPC, a 2-counter (one monitor and spectrometer) based system including a precision pump for sample injection and dilution. This system is able to count and size colloids from 50 nm to 2 mm into 19 size classes. Calibration work was performed with standard latex beads and natural samples from various surface waters. The quantification limit on the colloid number concentration for the smallest (50/100 nm), the medium (400/ 500 nm) and the largest (>2 mm) size classes are 1.0E4, 1.1E2 and 18 ml-1, respectively. Instrument saturation occurs at 8E8 ml-1 for the 50/100 size class, and at 2E3 ml-1 for medium and highest size classes. The relative repeatability varies between 7 and 29% for the different size classes in a synthetic, polydispersed sample reproducing typical environmental colloid concentration. These values are not significantly different from the with-in replicates variability. The verification of the factory calibration shows that measured concentrations are between 27 and 106% of the nominal concentration of standards. In conclusion, the presented SPC system gives reliable results of colloid number concentration and size distribution in a wide range of natural surface waters and offers new opportunities for on-site studies.