Multiple sclerosis (MS) is a chronic autoimmune disease of the central nervous system (CNS) characterized by inflammation, demyelination and neurodegeneration. The 18 kDa translocator protein (TSPO) is overexpressed on activated microglia, a key mediator of acute and chronic inflammation in MS. Positron emission tomography (PET) targeting TSPO enables in vivo detection of microglial activation in MS, revealing a high proportion of active white matter (WM) lesions. It also detects a substantial innate immune activation within normal-appearing white matter (NAWM), gray matter (GM), choroid plexus (CP), and meninges. In MS patients, increased TSPO binding correlates with clinical disability and brain atrophy, particularly of thalami and other deep GM structures, and predicts disease progression. TSPO PET also shows promise for monitoring responses to disease-modifying therapies (DMTs), and for detecting early signs of opportunistic infections, such as progressive multifocal leukoencephalopathy (PML). Integration with advanced MRI techniques and novel tracers beyond TSPO is likely to further improve our understanding of MS pathophysiology. Despite current limitations, including genetic polymorphisms affecting tracer binding and technical barriers to clinical implementation, TSPO PET may also represent a promising outcome measure for future treatments targeting microglia. Here, we critically review how PET targeting TSPO advances our understanding of MS pathophysiology, aids in prognostication, and may contribute to therapeutic monitoring.