ObjectivesTo introduce serialised medicines into an operational hospital dispensary and assess the technical effectiveness of digital medicine authentication (MA) technology under European Union Falsified Medicines Directive (EU FMD) conditions.DesignThirty medicine lines were serialised using 2D data matrix labels and introduced into an operational UK National Health Service (NHS) hospital dispensary. Staff were asked to check medicines for two-dimensional (2D) data matrices and scan those products, in addition to their usual medicine preparation and checking processes. Four per cent of the study medicines were labelled with a 2D barcode which generated a pop-up, identifying the medicine as either authenticated elsewhere (falsified), authenticated here, expired or recalled.SettingAn NHS teaching hospital based in the UK, the same site as the Naughton et al 2016 study.ParticipantsGeneral Pharmaceutical Council registered, accredited accuracy checking technicians and pharmacists.Primary outcome measuresAverage response times, offline issues, instances of incorrect quarantine and workarounds. The EU FMD maximum response time is 300 milliseconds (ms).ResultsDuring the checking stage of medicine preparation, the average response time for MA in this study was 131 ms. However, 4.67% of attempted authentications experienced offline issues, an increase of 4.23% from the previous study. An increase in offline instances existed alongside an increase in incorrect quarantine.ConclusionsDigital drug screening has the capability of operating with average response times which are below the maximum EU FMD limit of 300 ms. However, there was an increased incidence of offline errors and cases of incorrect quarantine. The practical and legal implications of supplying a substandard or falsified medicine during offline periods without prior authentication or withholding supply until online status resumes are not yet fully understood.