The approval of maraviroc (Selzentri®), the first CCR5 antagonist, with specific antiviral activity against CCR5 (R5)-tropic HIV variants, has promoted the determination of HIV coreceptor usage in the clinical setting. The phenotypic assay Trofile™, which is based on recombinant virus technology, has been the most widely used diagnostic test, given that it was the only assay which provided tropism information in the pivotal maraviroc clinical trials. However, this method displays logistical and technical limitations that make it far from convenient as a diagnostic test in clinical practice. Genotypic methods based on V3 genotyping represent a more feasible alternative and progressively are replacing phenotypic assays. Even though their sensitivity to detect X4-tropic variants is lower compared to Trofile™, recent studies have demonstrated that specific genotypic tools (geno2pheno and PSSM) are comparable to Trofile™ and ES-Trofile™ in predicting virologic response to maraviroc. This review summarizes clinical and methodological recommendations for the genotypic determination of HIV tropism to guide therapeutic decisions with CCR5 antagonists in HIV therapeutics.