Pre-exposure prophylaxis (PrEP) is recommended by the World Health Organization as an additional prevention option for people at substantial risk for HIV infection as part of combination HIV prevention. A key challenge when planning to implement PrEP is setting targets at national and subnational levels: how many individuals should a country or district aim to reach? This resource provides a method to estimate the number of individuals at substantial risk who may benefit from PrEP within specific key and high-priority populations. It includes technical materials describing the mathematical modelling that underpins the method, spreadsheets that can be used to develop estimates for five priority populations and instructions for using these tools.