Althoug frequency of occurrence (either as percentage or relative frequency) is the most common method of expressing the content of otter Lutra lutra faeces (spraints), the accuracy of the method, and the effects of varying sampling procedures (e.g. inter-collection interval) and sample sizes, have not been quantified. The validity of the technique was assessed in the present study by feeding trials involving captive, tame otters and computer simulation of various spraint sub-sampling regimes. Four animals were fed known quantities (numbers and biomass) of a total of nine fish species, two bird species and one mammal over a 28-day period. Most prey remains were passed in spraints within 24 h, although perch Perca fluviatilis scales appeared up to 10d after consumption. Remains from single meals of perch were recorded in 60 subsequent spraints from two otters, and remains of individual eels Anguillu anguilla were recorded in up to 11 spraints. Some single spraints contained the remains of up to seven individual salmonids, Salmo spp. Minnows Phoxinus phoxinus placed within the body cavities of larger rainbow trout Onrorhynchus mykiss were easily identified in spraints, as were the remains of Dytiscus spp. beetles which were not included in trial meals. The latter confirms that otters actively consumed large free-swimming insects. Spraint analysis accurately determined the rank order of prey groups for individual otters and for all four combined. However, few of the true proportions consumed fell within the 95% confidence limits calculated from spraints. Over the month-long trial, the overall picture of otter diet was altered little by increased inter-sampling period for spraints. But as samples were reduced, coefficients of variation for the mean estimates of each prey group increased and were often too large for estimates to be meaningful. It is not possible to quantify otter diet accurately by frequency of occurrence methods, and the results of previous studies attempting to quantify the amount of a specific prey item consumed by otters using this method must be treated with caution. Diet could be estimated more accurately from spraint analysis by concentrating on the main prey species and using keybones, which are resistant to digestion, to determine relative size-frequency distributions.