The diagnosis of rheumatoid arthritis (RA) must be made early, because prompt initiation of treatments tailored to disease activity is crucial to improve structural and functional outcomes. Anti-citrullinated peptide antibodies (ACPAs) are well-established diagnostic markers for RA and should be included in the classification criteria. Here, we describe the main tests for detecting ACPAs and we underline the diagnostic and prognostic usefulness of ACPAs in patients with RA. The presence of ACPAs predicts poorer functional and structural outcomes, and ACPA titers respond to some of the medications used in RA. Therefore, ACPA titers should be determined at regular intervals throughout follow-up.