The purpose of this document is to provide guidance on the use of pooled sample testing strategy in coronavirus disease (COVID-19) testing laboratories of the African Union Member States for scaling up SARS-CoV-2 nucleic acid testing capacity with the available resources. The current document describes the effect of factors such as the prevalence of COVID-19 in the population to be tested, the homogeneity of pools, and the sensitivity of the molecular test in optimal pool size determination. It also highlights the importance of monitoring the prevalence of COVID-19 in a population to be tested and proper validation of the test, to limit the potential for false-negative results. Validation studies to determine the optimal pool size by testing laboratories are recommended as the optimum approach. A safe, simple ‘two-stage pooling’ option has been indicated in this guidance to be used by laboratories until such validation can be achieved.