Polymerase chain reaction-based fluorescent Luminex assay to detect the presence of human papillomavirus types

Becuase 40% of human papillomavirus (HPV) infections are mixed infections, the accurate identification of high-risk HPV genotypes in mixed infections is important for defining a woman's risk for progression to cervical cancer. Thus, advanced Luminex-based HPV genotyping has been developed to simultaneously detect the presence of multiple HPV types. Here, we describe the development of a Luminex-based HPV genotyping that combines polymerase chain reaction amplification with hybridization to fluorescence-labeled polystyrene bead microarrays (Luminex suspension array technology). New HPV type-specific oligonucleotide probes and YBT L1/GP6-1 primers were used to detect the HPV types in 132 clinical samples. We simultaneously evaluated the usefulness of this technique on clinical samples. We detected 15 specific HPV types (6, 16, 18, 31, 35, 42, 51, 52, 55, 56, 58, 59, 66, 67 and 68) examined with specificity without known cross-reaction to other HPV types. The detection limit for the different HPV types was above 500 plasmids. We compared the performance of the Luminex-based assay to the established HPV DNA microarray chip for polymerase chain reaction products derived from 53 clinical samples. The evaluation showed excellent agreement. The Luminex-based HPV genotyping was a sensitive, reproducible technique for the simultaneous genotyping of all clinically relevant genital HPV types. This assay system may be used to provide critical clinical information for early detection of HPV, especially in cases where the HPV copy numbers are low and the latency period of HPV infection is prolonged.