Introduction: Misidentification of the extrahepatic bile duct anatomy during laparoscopic cholecystectomy (LC) is the main cause of bile duct injury. Easier intraoperative recognition of the biliary anatomy may be accomplished by using near-infrared fluorescence (NIRF) imaging after an intravenous injection of indocyanine green (ICG). Promising results were reported for successful intraoperative identification of the extrahepatic bile ducts compared to conventional laparoscopic imaging. However, routine use of ICG fluorescence laparoscopy has not gained wide clinical acceptance yet due to a lack of high-quality clinical data. Therefore, this multicentre randomised clinical study was designed to assess the potential added value of the NIRF imaging technique during LC. Methods and analysis: A multicentre, randomised controlled clinical trial will be carried out to assess the use of NIRF imaging in LC. In total, 308 patients scheduled for an elective LC will be included. These patients will be randomised into a NIRF imaging laparoscopic cholecystectomy (NIRF-LC) group and a conventional laparoscopic cholecystectomy (CLC) group. The primary end point is time to 'critical view of safety' (CVS). Secondary end points are 'time to identification of the cystic duct (CD), of the common bile duct, the transition of CD in the gallbladder and the transition of the cystic artery in the gallbladder, these all during dissection of CVS'; 'total surgical time'; 'intraoperative bile leakage from the gallbladder or cystic duct'; 'bile duct injury'; 'postoperative length of stay', 'complications due to the injected ICG'; 'conversion to open cholecystectomy'; 'postoperative complications (until 90 days postoperatively)' and 'cost-minimisation'. Ethics and dissemination: The protocol has been approved by the Medical Ethical Committee of Maastricht University Medical Center/Maastricht University; the trial has been registered at ClinicalTrials. gov. The findings of this study will be disseminated widely through peer-reviewed publications and conference presentations.