Centrifugal elutriation discriminates cells according to their sedimentation coefficient, generating homogeneous samples well suited for genomic comparative approaches. It can, for instance, isolate G1 daughter cells from a Saccharomyces cerevisiae unsynchronized population, alleviating aging and cell-cycle biases when conducting genome-wide/single-cells studies. The present report describes a straightforward and robust procedure to determine whether a cell population of virtually any yeast species can be efficiently elutriated, while offering solutions to optimize success. This approach was used to characterize elutriation parameters and S-phase progression of four yeast species (S. cerevisiae, Candida glabrata, Lachancea kluyveri and Pichia sorbitophila) and could theoretically be applied to any culture of single, individual cells. This article is protected by copyright. All rights reserved.