Xiaobin et al. BMC Cancer 2012, 12:340 http://www.biomedcentral.com/1471-2407/12/340 operative and postoperative management training for following the protocol. The abdominal incision can be decided according to the surgeon’s preference. A complete exploration of the abdomen and an intraopera- tive ultrasound evaluation of the whole liver will be per- formed to decide whether curative resection is feasible. If necessary, frozen section biopsies will be taken to evaluate suspicious lesions or lymph nodes. If both inter- vention procedures seem possible to the surgeon, randomization will be performed at this time point. Experimental group (group a, non-Pringle group) The liver will be mobilized from the retroperitoneum and the peripheral ligament. Anatomical liver resection is preferable. If anatomical resection is not possible, a surgical margin of more than 2 cm should be achieved, except if the location of tumor nodes makes this impos- sible (e.g., close to the inferior vena cava or portal vein). The resection method and surgical margin will be recorded for analysis. The liver parenchyma can be resected according to the surgeon’s preference and local standards (CUSA, Tissue-link.). The Pringle maneuver will not be used, and hepatic inflow and outflow will be maintained. For safety, the surgeon may use the Pringle maneuver if major bleeding occurs. Control group (group B, Pringle maneuver group) After mobilization, the Pringle maneuver will be per- formed. The same approach will be used for hepatic par- enchymal transection as in the experimental group. If necessary, the outflow of the hepatic vein can be con- trolled, and this factor will be analyzed. For safety, the surgeon may change the surgical technique at any time during the operation. Pringle maneuver The portal triad clamping is set to 15 min hepatic inflow occlusion followed by 5 min of reperfusion, repeated as needed. The last occlusion should be 15 min even if the transection has been completed. The total ischemia and reperfusion times and the number of occlusion cycles will be recorded for analysis. Any deviation in the stand- ard operating procedures will be regarded as a protocol violation. Surgical team The surgical teams consist of senior surgeons who all have standard qualifications and have independently per- formed standard anatomical hepatectomy in more than 100 patients. The results of randomization will be made known to the operating surgeon only after the disease has been deemed suitable for curative resection. All sur- gical procedures and anesthesia will be performed by the specifically trained teams of experienced hepatobiliary Page 4 of 8 surgeons and anesthesiologists, ensuring standardized execution of the study protocol. Patients will be ran- domly assigned to eligible surgeons to minimize the effects of variations between surgeons on operative outcomes. Preoperative examination Preoperative examination of patients will include blood biochemistry, alpha-fetoprotein assay, chest x-ray, percu- taneous ultrasonography, computed tomography (CT) scan, and hepatic angiography in selected patients. Liver function will be assessed by Child-Pugh grading and the indocyanine green clearance test. Acquisition of samples Blood samples: Two blood samples will be obtained from each patient after induction of general anesthesia through a central venous catheter, which is routinely placed just before surgery. EDTA (not heparin) will be used as an anticoagulant for blood samples. Follow-up blood samples will be collected every 6 months postoperatively. Tissue samples: Paracarcinomatous tissue and cancer tissue will be harvested from the resected liver specimen at the time of resection. Paracarcinomatous tissue should be harvested 1 cm from the tumor margin. All tumors will be pathologically confirmed to be HCC. Each tissue sample will be divided into two parts, one of which will be placed in liquid nitrogen and the other in 10% formalin. Serial 4-μm sections from each speci- men will be stained with hematoxylin and eosin to deter- mine clinicopathological features such as venous invasion, capsule formation, Edmondson’s grade, and cir- rhotic nodules. Sample size calculation and data analysis plan The sample size of this trial was calculated using data from our previous experience and from the published literature. There are currently no data from randomized controlled trials showing the recurrence rate of HCC after hepatectomy without the Pringle maneuver. Most studies used the Pringle maneuver. This trial plans to have equal numbers of control and experimental sub- jects, and a 36-month follow-up period. The median sur- vival time after the treatment that the control group will receive has been reported to be 26 months [24,44-50]. We anticipate a10-month difference in the median recurrence-free survival time between the experimental and control groups. If median survival times in the con- trol and experimental groups are 26 months and 36 months, respectively, we will need to include 249 ex- perimental subjects and 249 control subjects to be able to reject the hypothesis that the experimental and control survival curves are equal with a probability