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Pooling of samples can increase throughput and reduce costs for large-scale SARS-CoV-2 testing when incidence is low. In a cross-sectional study of serial SARS-CoV-2 sampling of staff and residents at three nursing homes, laboratory labor constraints limited the feasibility of pooling prior to the maximal incidence that favored cost savings. IMPORTANCE This study highlights the pragmatic considerations surrounding SARS-CoV-2 sample pooling beyond accuracy and costs. We performed a cost analysis to determine the percent positivity at which pooling would reduce costs versus single testing. We found that the need for a stable amount of daily work hours staffed by a highly trained workforce was a major limitation in pooling as test positivity increased. For the COVID-19 pandemic and future pandemic threats, laboratories should carefully consider the thresholds at which sample pooling is beneficial, with a particular focus on the impact on laboratory staff.