title: Next generation digital PCR measurement of hepatitis B virus copy number in formalin-fixed paraffin-embedded hepatocellular carcinoma tissue.
abstract: BACKGROUND
  Hepatocellular carcinoma (HCC) is strongly associated with hepatitis B virus (HBV) infection. False-negative results are common in routine serological tests and quantitative real-time PCR because of HBV surface antigen (HBsAg) variation and low HBV copy number. Droplet digital PCR (ddPCR), a next generation digital PCR, is a novel, sensitive, and specific platform that can be used to improve HBV detection.
  
  
  METHODS
  A total of 131 HCC cases with different tumor stages and clinical features were initially classified with a serological test as HBsAg positive (n = 107) or negative (n = 24) for HBV infection. Next, DNA templates were prepared from the corresponding formalin-fixed paraffin-embedded (FFPE) tissues to determine HBV copy number by ddPCR.
  
  
  RESULTS
  HBV copy numbers, successfully determined for all clinical FFPE tissues (n = 131), ranged from 1.1 to 175.5 copies/μL according to ddPCR. The copy numbers of HBV were positively correlated with tumor-nodes-metastasis (P = 0.008) and Barcelona-Clinic Liver Cancer (P = 0.045) classification. Moreover, serum cholinesterase correlated with hepatitis B viral load (P = 0.006).
  
  
  CONCLUSIONS
  HBV infection is a key factor that influences tumorigenesis in HCC by regulating tumor occurrence and development. ddPCR improves the analytical sensitivity and specificity of measurements in nucleic acids at a single-molecule level and is suitable for HBV detection.
authors.affiliation: Center for Gene Diagnosis.
authors.forename: Jing-Tao
authors.initials: JT
authors.lastname: Huang
authors.affiliation: Medical Research Center, and.
authors.forename: Ying-Juan
authors.initials: YJ
authors.lastname: Liu
authors.affiliation: Department of Translational Molecular Pathology, The University of Texas MD Anderson Cancer Center, Houston, TX;
authors.forename: Jin
authors.initials: J
authors.lastname: Wang
authors.affiliation: Department of Pathology, Zhongnan Hospital of Wuhan University, Wuhan, China;
authors.forename: Zhi-Gao
authors.initials: ZG
authors.lastname: Xu
authors.affiliation: Center for Gene Diagnosis.
authors.forename: Ying
authors.initials: Y
authors.lastname: Yang
authors.affiliation: Center for Gene Diagnosis.
authors.forename: Fan
authors.initials: F
authors.lastname: Shen
authors.affiliation: Department of Clinical Laboratory, Gongli Hospital, Second Military Medicine University, Shanghai, China.
authors.forename: Xing-Hui
authors.initials: XH
authors.lastname: Liu
authors.affiliation: Center for Gene Diagnosis.
authors.forename: Xin
authors.initials: X
authors.lastname: Zhou
authors.affiliation: Center for Gene Diagnosis, Medical Research Center, and smliu@whu.edu.cn.
authors.forename: Song-Mei
authors.initials: SM
authors.lastname: Liu
mesh_terms: D006528:Carcinoma, Hepatocellular / Q000473:pathology / Q000821:virology*; D004270:DNA, Circular / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics; D005260:Female; D018628:Gene Dosage; D006509:Hepatitis B / Q000473:pathology / Q000821:virology*; D006514:Hepatitis B Surface Antigens / Q000097:blood / Q000235:genetics; D006515:Hepatitis B virus / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D008113:Liver Neoplasms / Q000473:pathology / Q000821:virology*; D008297:Male; D008875:Middle Aged; D009367:Neoplasm Staging; D016612:Paraffin Embedding; D016133:Polymerase Chain Reaction / Q000379:methods
chemical_list: D004270:DNA, Circular; D004279:DNA, Viral; D006514:Hepatitis B Surface Antigens