Molecular imaging tests frequently are performed as part of the diagnostic workup of musculoskeletal infection. Three-phase bone scintigraphy reliably diagnoses osteomyelitis in bones not affected by underlying conditions. The test is less useful, because of decreased specificity, in patients with underlying bony abnormalities or alterations such as fractures, orthopaedic hardware, arthritic changes, and tumors. At one time gallium-67 scintigraphy was used as a complement to bone scintigraphy to improve the specificity of diagnosis. With the introduction of labeled leukocytes and, more recently, fluorine-18 fluorodeoxyglucose, gallium-67 is reserved primarily for spinal infections when fluorine-18 fluorodeoxyglucose is not available. Except for the spine, in vitro labeled leukocyte imaging is valuable for diagnosing osteomyelitis in the setting of underlying osseous abnormalities. Leukocytes accumulate in bone marrow as well as in the presence of infection. Performing complementary bone marrow imaging with technetium-99m sulfur colloid facilitates the differentiation between the two and improves test accuracy. Fluorine-18 fluorodeoxyglucose accurately diagnoses spondylodiskitis and diabetic foot osteomyelitis, but its value in periprosthetic joint infection has yet to be determined.