This document was developed to provide practical guidance on interpretation of the most commonly used first- and second-line line probe assays (LPAs). In this updated manual, the interpretation of mutations identified by the assays has been revised to align with the WHO catalogue of mutations in Mycobacterium tuberculosis complex and their association with drug resistance. The latest LPA instructions for use of the assays are also presented. Lastly, case studies are provided that support staff at national and regional TB reference laboratories to understand and manage any discrepancies between phenotypic and genotypic drug susceptibility testing results