Previous studies have implicated viral infections in the pathogenesis of sudden unexpected death in infancy (SUDI), and routine virological investigations are recommended by current SUDI autopsy protocols. The aim of this study is to determine the role of post-mortem virology in establishing a cause of death. A retrospective review of 546 SUDI autopsies was carried out as part of a larger series of >1,500 consecutive paediatric autopsies performed over a 10-year period, 1996-2005, in a single specialist centre. Virological tests were performed as part of the post-mortem examination in 490 (90%) of the 546 SUDI autopsies, comprising 4,639 individual virological tests, of which 79% were performed on lung tissue samples. Diagnostic methods included immunofluorescence assays (using a routine respiratory virus panel; 98% of cases), cell culture (61%), rapid culture techniques such as the DEAFF test for CMV (55%), PCR (13%), electron microscopy (10%), and others. Virus was identified in only 18 cases (4%), viz. five cases of enterovirus, four of RSV, three of HSV and CMV, and one each of adenovirus, influenza virus and HIV. In seven of the 18 cases the death was classified as due to viral infection, whilst of the remaining 11 cases, death was due to bacterial infection in five, a non-infective cause in one and unexplained in five. Virus was identified in 33% of deaths due to probable viral infections, but also in 6% of SUDI due to bacterial infections, and in 2% of SUDI due to known non-infective causes and unexplained SUDI. When predominantly using immunofluorescence, virus is identified in only a small proportion of SUDI autopsies, resulting in a contribution to the final cause of death in <2% of SUDI post-mortem examinations. Routine post-mortem virological analysis by means of an immunofluorescence respiratory virus panel appears to be of limited benefit in SUDI for the purposes of determining cause of death. Application of a broader panel using more sensitive detection techniques may reveal more viruses, although their contribution to the final cause of death requires further exploration.