In the last two decades, the development of high-throughput diagnostic methods and the availability of effective treatments have increased the interest in newborn screening for lysosomal storage disorders. However, long-term follow-up experience is needed to clearly identify risks, benefits and challenges. We report our 8-year experience of screening and follow-up on about 250,000 neonates screened for four lysosomal storage diseases (Pompe disease, mucopolysaccharidosis type I, Fabry disease, Gaucher disease), using the enzyme activity assay by tandem mass spectrometry, and biomarker quantification as a second-tier test. Among the 126 positive newborns (0.051%), 51 infants were confirmed as affected (positive predictive value 40%), with an overall incidence of 1:4874. Of these, three patients with infantile-onset Pompe disease, two with neonatal-onset Gaucher disease and four with mucopolysaccharidosis type I were immediately treated. Furthermore, another four Gaucher disease patients needed treatment in the first years of life. Our study demonstrates the feasibility and effectiveness of newborn screening for lysosomal storage diseases. Early diagnosis and treatment allow the achievement of better patient outcomes. Challenges such as false-positive rates, the diagnosis of variants of uncertain significance or late-onset forms and the lack of treatment for neuronopathic forms, should be addressed.