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Environmental DNA (eDNA) sampling is a promising method for surveying aquatic fauna. Recent eDNA studies have investigated the likelihood of false negative errors in the laboratory and in the field, but the likelihood of false positives remains poorly studied. We investigated the likelihood of both types of errors in eDNA surveys of an Australian threatened freshwater fish (Galaxiella pusilla) using laboratory experiments, field surveys, and recent advances in hierarchical site occupancy‐detection modeling. Laboratory experiments revealed high primer/probe specificity; absence of sample contamination in extraction and qPCR blanks; and rapid accumulation and deterioration of eDNA in aquaria. Hierarchical site occupancy‐detection models fitted to pilot data collected at 13 wetlands revealed that two water samples, each with two qPCRs, would be required to achieve a cumulative detection probability >.95. A …