The immunocompromised host is a particularly vulnerable population in whom routine and unusual infections can easily and frequently occur. Prosthetic devices are commonly used in these patients and the infections associated with those devices present a number of challenges for both the microbiologist and the clinician. Biofilms play a major role in device-related infections, which may contribute to failed attempts to recover organisms from routine culture methods. Moreover, device-related microorganisms can be difficult to eradicate by antibiotic therapy alone. Changes in clinical practice and advances in laboratory diagnostics have provided significant improvements in the detection and accurate diagnosis of device-related infections. Disruption of the bacterial biofilm plays an essential role in recovering the causative agent in culture. Various culture and nucleic acid amplification techniques are more accurate to guide directed treatment regimens. This chapter reviews the performance characteristics of currently available diagnostic assays and summarizes published guidelines, where available, for addressing suspected infected prosthetic devices.