title: Comparison of Droplet Digital PCR and Real Time PCR Method for HBV DNA Quantification.
abstract: BACKGROUND
  HBV infection causes a potential serious public health problem. The ability to detect HBV DNA concentration is an important issue that had been continuously improved. When using quantitative polymerase chain reaction (qPCR), several factors are of concern, for example, sources of material, standard curve calibration, and PCR efficiency. Digital PCR (dPCR) is an alternative PCR-based technique for absolute quantification using Poisson's statistics without requiring a standard curve.
  
  
  OBJECTIVE
  Compare the data set of HBV DNA generated between dPCR and qPCR methods.
  
  
  METHODS
  Fifty-four samples were quantified by Abbot's real time PCR and with 2-6 log10 HBV DNA were selected for comparison with dPCR.
  
  
  RESULTS
  Of these 54 samples, there were two outlier samples defined as negative by dPCR, whereas 52 samples were positive by both of these assays. The difference between two assays was less than 0.25 log IU/mL in 24/52 samples (46%) of paired samples; less than 0.5 log IU/mL in 46/52 samples (88%) and less than 1 log in 50/52 samples (96%). The correlation coefficient (r) was 0.788 (p-value < 0.0001). Comparison with qPCR method, data generated by dPCR tend to be an overestimation in the sample with the low level ofHBVDNA concentration and underestimated in the sample with high viral load. The variation of DNA by dPCR measurement might be due to the pre-amplification procedure and PCR template.
  
  
  CONCLUSIONS
  Measurement of HBV DNA by using dPCR, the results ofthe HBV DNA copy number tended to be deviated by over- or under-estimated when comparison to real time PCR method. In addition, a large quantity of DNA was used when compared to qPCR. However, the optimum processes of this assay have to be further investigated.
authors.forename: Chatchawal
authors.initials: C
authors.lastname: Wongjitrat
authors.forename: Navin
authors.initials: N
authors.lastname: Horthongkham
authors.forename: Ruengpung
authors.initials: R
authors.lastname: Sutthent
authors.forename: Surangrat
authors.initials: S
authors.lastname: Srisurapanon
mesh_terms: D002138:Calibration; D004279:DNA, Viral / Q000097:blood*; D006509:Hepatitis B / Q000175:diagnosis*; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D019562:Viral Load
chemical_list: D004279:DNA, Viral