This study aimed to investigate the effects of two faecal collection methods (stripping and settlement) on the apparent digestibility coefficients (ADC) of dry matter, protein and energy of three different diets fed to barramundi. In a second experiment, the effect of acclimation time (i.e. number of days fed the diet) on the calculation of ADCs was also investigated. Each tank of fish was fed one of three diets for 12 days. Faeces were collected by both stripping and settlement, although only settlement was used prior to day seven of the acclimation period. Faeces were collected using the settlement method at regular intervals from day 1 to day 12. Comparisons between faecal collection methods were only made based on faecal material collected over a similar acclimation period. The collection of faeces by stripping produced more conservative ADCs, which were also more consistent than those obtained using the settlement technique. The calculated ADCs typically fluctuated for the first 3 days of collection before the variability diminished. Barramundi should be acclimated to diets for a minimum of 4 days before collection of faecal material, and collection by stripping is recommended to obtain the most reliable digestibility data.