This study examines the utility of resistance testing during treatment interruption, making comparisons with the current IAS guidelines. A total of 188/1279 tests (14%) were performed during treatment interruption; 69/188 tests (36.7%) demonstrated key mutations. Time off therapy and the total number of previous drugs were both significantly associated with the presence of mutations. We conclude that resistance testing is of value up to 3 months after treatment interruption, and may convey some benefit up to 12 months.