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Open Access Methodology

A new in situ hybridization and immunohistochemistry with a novel antibody to detect small T-antigen expressions of Merkel cell polyomavirus (MCPyV)

Michiko Matsushita12, Daisuke Nonaka3, Takeshi Iwasaki1, Satoshi Kuwamoto1, Ichiro Murakami1, Masako Kato1, Keiko Nagata1, Yukisato Kitamura2 and Kazuhiko Hayashi1*

Author Affiliations

1 Division of Molecular Pathology, Department of Pathology, Tottori University Faculty of Medicine, Yonago, Japan

2 Department of Pathobiological Science and Technology, School of Health Science, Tottori University Faculty of Medicine, Yonago, Japan

3 Department of Histopathology, The Christie NHS Foundation Trust, Manchester, UK

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Diagnostic Pathology 2014, 9:65  doi:10.1186/1746-1596-9-65

Published: 20 March 2014

Abstract

Background

Approximately 80% of Merkel cell carcinomas (MCCs) harbor Merkel cell polyomavirus (MCPyV) which monoclonally integrates into the genome and has prognostic significance. The presence or absence of MCPyV is usually diagnosed using CM2B4 immunohistochemistry (IHC) for MCPyV-large T antigen (LT) protein. However, this method poses a risk of misdiagnosis.

Methods

In this study, we determined MCPyV infection in MCCs using real-time PCR for MCPyV-LT DNA and prepared 16 cases of MCPyV-DNA-positive and -negative groups. Diagnostic sensitivity and specificity of conventional PCR for MCPyV-small T antigen (MCPyV-ST), IHC using a newly developed polyclonal antibody (ST-1) for MCPyV-ST protein (MCPyV-ST) (aa: 164–177), and in situ hybridization (ISH) as well as real-time PCR for MCPyV-ST mRNA were compared against CM2B4-IHC for sensitivity (0.94, 15/16) and specificity (0.94, 15/16).

Results

The followings are the respective sensitivity and specificity results from examinations for MCPyV-ST gene: conventional PCR for the MCPyV-ST (0.94, 1.0), ST-1-IHC (0.69, 1.0), real-time PCR for ST mRNA (1.0, no data), ST mRNA ISH (0.94, 1.0). Each of the MCPyV-pseudonegative (1/16) and -pseudopositive (1/16) diagnoses evaluated using CM2B4-IHC were accurately corrected by examinations for MCPyV-ST or its expression as well as real-time PCR for MCPyV-LT. Sensitivity of CM2B4-IHC (0.94) was superior to that of ST-1-IHC (0.69) but equal to that of ST mRNA-ISH (0.94). Specificities of ST-1-IHC (1.0) and ST mRNA-ISH (1.0) were superior to that of CM2B4-IHC (0.94).

Conclusions

Therefore, combined application of ST mRNA-ISH and ST-IHC as well as CM2B4-IHC is recommended and will contribute to the diagnostic accuracy for MCPyV infection in MCCs.

Virtual slides

The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/9966295741144834 webcite

Keywords:
Merkel cell carcinoma; MCPyV-small T antigen; ST-immunohistochemistry; ST mRNA-ISH