Utility of p16INK4a staining on cell blocks prepared from residual liquid-based cervicovaginal material.

Abstract

Introduction: Cervical cancer screening using Pap smears is affected by false-negative results. Liquid-based cytology (LBC) offers the technical advantage of preparing cell blocks from residual fluid to conduct ancillary tests on them. The p16INK4a gene product has been shown to be strongly overexpressed in dysplastic cervical epithelia and serves as surrogate marker for high-risk human papilloma virus (HPV) infection.

Material and methods: Microwave-processed cell blocks were prepared from residual material in vials after ThinPrep slide preparation, stained with hematoxylin and eosin (H&E) and p16INK4a. Nuclear staining with or without cytoplasmic staining on p16 slides was considered positive. Four parameters were evaluated: percentage of positive cells, intensity of staining, number of positively stained cells in close contact and full-thickness epithelial staining. We compared sensitivity and specificity of ThinPrep smears and p16 stained cell blocks in diagnosing invasive malignancy.

Results: The intensity and percentage of p16 positive cells was found to increase with increasing grade of cervical abnormality. We found good concordance between ThinPrep smear and cell block diagnoses in cases which were negative for intraepithelial lesion or malignancy (NILM) (97.6%), in low-grade squamous intraepithelial lesions (LSIL) (90%), high-grade squamous intraepithelial lesions (HSIL) (100%) and squamous cell carcinomas (SCC) (93.5%). Of 16 discrepant cases, nine were reported unsatisfactory on ThinPrep smears due to abundant necrosis or scant cellularity. All these turned out to have malignancies on follow up and review of histology. The sensitivity of ThinPrep and p16 stained cell blocks in diagnosing invasive malignancy were 70.2% and 85.1% respectively, while the specificity of both were 100%.

Conclusions: Cell blocks prepared from residual fluid in LBC vials have the potential to reduce the rates of inadequacy and are feasible in routine practice. While the cost of p16 on cell blocks may be too prohibitive for use in routine cervical screening programmes, if used judiciously in combination with clinical suspicion, a lot of valuable material which is usually discarded in the residual LBC vials can prove to be crucial in arriving at the correct diagnosis.