Through sample-size-based rarefaction analyses, we tried to suggest the appropriate degree of sample concentration and sub-sample extraction, as a way to estimate more accurate zooplankton species diversity when assessing biodiversity. When we collected zooplankton from three reservoirs with different environmental characteristics, the estimated species richness (S) and Shannon’s Hʹ values showed different changing patterns according to the amount of sub-sample extracted from the whole sample by reservoir. However, consequently, their zooplankton diversity indices were estimated the highest values when analyzed by extracting the largest amount of sub-sample. As a result of rarefaction analysis about sample coverage, in the case of deep eutrophic reservoir (Juam) with high zooplankton species and individual numbers, it was analyzed that 99.8% of the whole samples were represented by only 1 mL of sub-sample based on 100 mL of concentrated samples. On the other hand, in Soyang reservoir, which showed very small species and individual numbers, a relatively low representation at 97% when 10 mL of sub-sample was extracted from the same amount of concentrated sample. As such, the representation of sub-sample for the whole zooplankton sample varies depending on the individual density in the sample collected from the field. If the degree of concentration of samples and the amount of subsample extraction are adjusted according to the collected individual density, it is believed that errors that occur when comparing the number of species and diversity indices among different water bodies can be minimized.