Measuring transmission of a vector-borne infection is essential to understanding infection dynamics. When infection prevalence in the vector population is low, transmission is often measured by pool screening (also referred to as group testing). Several investigators have developed statistical methods to recover infection prevalence estimates from pool screen data. These are based on models that contain certain assumptions, and a pool screening approach must be designed to take these into account if accurate estimates of infection prevalence are to be obtained. Here we describe these assumptions and discuss appropriate sampling protocols. The sources of error inherent in pool screening are described, and we show that, under most conditions in which one would want to use group testing, most of the error results from sampling and not the pooling process. Issues involved in developing a sampling protocol, including the total number of insects to be screened and optimal pool size, are explored. The meaning of confidence intervals associated with prevalence estimates and the appropriate interpretation of these intervals are discussed. Copyright © 2006 by The American Society of Tropical Medicine and Hygiene.