Laboratory diagnostics are indispensable to modern healthcare, yet the integrity of test results can be compromised by medication-related interference (MRI), a significant and underappreciated threat to patient safety. Erroneous laboratory data stemming from MRI can precipitate a cascade of clinical errors, including misdiagnosis, delayed or inappropriate treatment, and unnecessary additional testing. This review offers a practical, evidence-based guide for the key professionals’ integral to the laboratory testing process: nursing technicians, clinical pharmacists, and laboratory specialists. We systematically explore the foundational mechanisms of interference, classifying them into physiological (in vivo) effects, where a drug's pharmacodynamic or toxic action genuinely alters analyte levels, and analytical (in vitro) effects, where a drug or its metabolite directly disrupts the assay methodology. The review examines the vulnerabilities of common analytical platforms—spectrophotometry, immunoassays, enzymatic methods, and chromatography—to such interferences. A substantial compendium details clinically significant MRIs caused by major drug classes, with a particular focus on the profound and dangerous impact of high-dose biotin supplementation on immunoassays. The central aim of this guide is to delineate the distinct yet synergistic roles of the multidisciplinary team. It highlights the nursing technician's pivotal role in the pre-analytical phase, the clinical pharmacist's expertise in proactive drug identification and management, and the laboratory specialist's critical function in result validation and interference investigation. By elucidating the complexities of MRI and championing a framework for robust interprofessional collaboration, this guide seeks to arm healthcare teams with the knowledge and strategies required to protect the integrity of laboratory data, thereby enhancing clinical decision-making and safeguarding patient safety.