Evaluation of quantity and staining pattern of human papillomavirus (HPV)-infected epithelial cells in thin-layer cervical specimens using optimized HPV-CARD assay

BACKGROUND. Testing for human papillomavirus (HPV) is used in the triage of women with a cervical cytology of atypical squamous cells of undetermined significance (ASCUS). A fluorescent in situ hybridization assay was developed for the detection of HPV using the catalyzed receptor deposition technique (HPV-CARD). In this study, the utility of this assay was tested for the detection of HPV in liquid-based cervical cytology specimens. METHODS. A total of 195 liquid-based cytology specimens were analyzed using the HPV-CARD assay. The results from the assay were compared with HPV polymerase chain reaction (PCR) and typing results. The number of HPV-infected cells and the staining pattern was correlated with the cytology classification. RESULTS. A 91% concordance between HPV-CARD and PCR was observed for the detection of high-risk HPV viruses. A 78% concordance was observed for specimens that were negative for HPV. In ASCUS, low-grade squamous intraepithelial lesion (LSIL), and high-grade squamous intraepithelial lesion (HSIL) categories, the average number of HPV-positive cells per slide was 19 cells, 127 cells, and 450 cells, respectively. The number of cells with a punctate staining, suggestive of HPV integration, was 21% in ASCUS, 34% in LSIL, and 46% in HSIL specimens. CONCLUSIONS. The results of the current study indicate positive correlations between the severity of the disease and the increased overall quantity of HPV-positive epithelial cells in cervical cytology specimens and accumulation of cells with punctate staining suggestive of integrated HPV. In summary; the developed HPV-CARD assay was found to provide novel information regarding the proportion and staining pattern of HPV-infected epithelial cells in different cytologic categories of cervical specimens.