title: Combined SYBR Green real-time polymerase chain reaction and microarray method for the simultaneous determination of human papillomavirus loads and genotypes.
abstract: OBJECTIVE
  The aim of this study was to describe the principle of the Cheil HPV DNA Chip assay and evaluate its accuracy. In order to quantify the human papillomavirus (HPV) load and identify HPV genotypes simultaneously, this assay combined the two methods: SYBR Green quantitative real-time polymerase chain reaction (PCR) and DNA microarray.
  
  
  METHODS
  We designed novel consensus primer sets that target the conserved region of the HPV L1 gene for quantifying and detecting a broad range of HPV types by quantitative real-time PCR. Subsequently, using the PCR products, DNA microarray was performed with 36 HPV type-specific probes. To validate this method, direct sequencing and correlation analysis among HPV genotype, viral load, and cytological abnormality was performed by Cohen's kappa values, two-sided McNemar chi-square test, Kruskal-Wallis test, and odds ratios.
  
  
  RESULTS
  The kappa value of the Cheil HPV DNA Chip was 0.963 (95% confidence interval, 0.919 to 0.98), which was significantly higher than the value of 0.527 (95% confidence interval, 0.447 to 0.59) obtained using a conventional HPV DNA Chip. HPV16 (χ2=62.28,P<0.01), HPV33 (χ2=7.18,P<0.01), and HPV58 (χ2=9.52,P<0.01), which are classified as high-risk HPVs, were detected at significant levels in samples with high-grade lesions. And viral loads tended to be higher in groups with high odds ratios.
  
  
  CONCLUSIONS
  The Cheil HPV DNA Chip is an effective diagnostic assay for simultaneously detecting HPV genotypes and loads in cervical samples.
authors.affiliation: Laboratory of Research and Development for Genomics, Cheil General Hospital and Women's Healthcare Center, Dankook University College of Medicine, Seoul, Korea.
authors.forename: Hyun Hee
authors.initials: HH
authors.lastname: Seo
authors.affiliation: Laboratory of Research and Development for Genomics, Cheil General Hospital and Women's Healthcare Center, Dankook University College of Medicine, Seoul, Korea.
authors.forename: Young Jun
authors.initials: YJ
authors.lastname: Kim
authors.affiliation: Laboratory of Research and Development for Genomics, Cheil General Hospital and Women's Healthcare Center, Dankook University College of Medicine, Seoul, Korea.
authors.forename: Mi Seon
authors.initials: MS
authors.lastname: Jeong
authors.affiliation: Department of Pathology, Cheil General Hospital and Women's Healthcare Center, Dankook University College of Medicine, Seoul, Korea.
authors.forename: Sung Ran
authors.initials: SR
authors.lastname: Hong
authors.affiliation: Department of Obstetrics and Gynecology, Cheil General Hospital and Women's Healthcare Center, Dankook University College of Medicine, Seoul, Korea.
authors.forename: In Ho
authors.initials: IH
authors.lastname: Lee
authors.affiliation: Department of Obstetrics and Gynecology, Cheil General Hospital and Women's Healthcare Center, Dankook University College of Medicine, Seoul, Korea.
authors.forename: Kyeong A
authors.initials: KA
authors.lastname: So
authors.affiliation: Department of Obstetrics and Gynecology, Cheil General Hospital and Women's Healthcare Center, Dankook University College of Medicine, Seoul, Korea.
authors.forename: Mi-Kyung
authors.initials: MK
authors.lastname: Kim
authors.affiliation: Department of Obstetrics and Gynecology, Cheil General Hospital and Women's Healthcare Center, Dankook University College of Medicine, Seoul, Korea.
authors.forename: Yoo Kyung
authors.initials: YK
authors.lastname: Lee
authors.affiliation: Department of Obstetrics and Gynecology, Cheil General Hospital and Women's Healthcare Center, Dankook University College of Medicine, Seoul, Korea.
authors.forename: Ki Heon
authors.initials: KH
authors.lastname: Lee
authors.affiliation: Department of Radiation Oncology, Cheil General Hospital and Women's Healthcare Center, Dankook University College of Medicine, Seoul, Korea.
authors.forename: Juree
authors.initials: J
authors.lastname: Kim
authors.affiliation: Department of Anatomy and Cell Biology, Hanyang University College of Medicine, Seoul, Korea.
authors.forename: Sung Jae
authors.initials: SJ
authors.lastname: Kim
authors.affiliation: Department of Obstetrics and Gynecology, Cheil General Hospital and Women's Healthcare Center, Dankook University College of Medicine, Seoul, Korea.
authors.forename: Tae Jin
authors.identifier: 0000-0002-5322-2745
authors.initials: TJ
authors.lastname: Kim
keywords: Diagnosis; Genotyping; Human papilloma virus; Microarray; Viral load