A Monoclonal-Antibody-Based Immunohistochemical Method for the Detection of Swine Influenza Virus in Formalin-Fixed, Paraffin-Embedded Tissues

Abstract

Swine influenza virus (SIV) is one of the most common respiratory viruses affecting swine. Swine influenza is present worldwide, and in the USA approximately 25-33% of slaughter pigs (6-7 months old) and 45% of 2-year-old breeding stock have anti-H1N1 SIV antibodies. Nearly all swine influenza in the USA is caused by H1N1 viruses. Only H1N1 viruses have been isolated from swine in the USA, although serologic studies indicate a low prevalence of antibody to H3N2 influenza virus. In 1994, infection with SIV was detected by virus isolation (VI), by fluorescent antibody examination (FA), or by recognition of characteristic lesions in 180 of approximately 1,800 cases of swine respiratory disease submitted to Iowa State University Veterinary Diagnostic Laboratory (ISU-VDL). Rapid, reliable methods for the diagnosis of influenza in swine are essential for veterinary diagnostic laboratories serving the swine industry. Currently, diagnosis is based on characteristic clinical signs, gross and microscopic lesions, results of FA examination of frozen tissue sections, and results of VI. Each diagnostic method has certain disadvantages. Clinical signs and gross and microscopic lesions may be atypical. The multifocal distribution of SIV lesions may result in sample-to-sample variation, which can affect the results of histopathologic and FA examinations. Other factors limiting the effectiveness of the FA test in demonstrating SIV antigen include the brief time postinfection that virus is present in the lung and the need to have fresh tissues with minimal autolytic changes available for examinations. In addition, use of a fluorescence microscope is necessary, and the stained slides are not permanent records because fluorescence fades quickly. The immunohistochemical (IHC) test detects virus in formalin-fixed, paraffin-embedded tissues and is relatively rapid, inexpensive, and easy to perform. The location of the virus in airway epithelial cells, macrophages, and pneumocytes can be directly visualized and associated with characteristic lesions by light microscopy. The IHC test also may be useful for performing retrospective studies when fresh tissue may no longer be available, because antigens detected by this method have been shown to be stable in paraffin for 10 years. The purpose of this study was to develop an IHC test for detection of SIV in formalin-fixed tissue. A similar