Monoclonal and Polyclonal Antibodies for Immunohistochemical Detection of Bovine Parainfluenza Type 3 Virus in Frozen and Formalin-Fixed Paraffin-Embedded Tissues

Abstract

Abstract. Accurate identification of bovine Parainfluenza type 3 virus in bovine respiratory disease requires dependable, sensitive, and specific techniques for detection in affected animals. Immunohistochemical testing can be a rapid and reliable means of demonstration of virus in tissues from suspect cases; however, this procedure is dependent upon the quality of the antisera directed against the viral antigens. The production of rabbit polyclonal and murine monoclonal antibodies directed against bovine Parainfluenza type 3 virus and techniques for their use in fresh-frozen and formalin-fixed paraffin-embedded tissues in immunofluorescence and immunoperoxidase-based immunohistochemical tests are described.

Immunohistochemical Detection of Parainfluenza Type 3 Virus Antigens in Paraffin Sections of Pneumonic Caprine Lungs

“…In rats experimentally infected with bovine PI-3 virus, viral antigens were present in the bronchial, bronchiolar and alveolar epithelial cells (Breker-Klassen et al, 1995). In the present study, the staining pattern, localization and distribution of antigens of the PI-3 virus antigens were almost similar to those described by Haines et al (1992) and Breker-Klassen et al (1995). Additionally, viral antigens were also demonstrated within exudates in airway lumina, epithelial cells of bronchial glands, multinucleated cells within alveolar lumina, lymphoid cells around vessels, the chondrocytes of cartilage around of small bronchi.…”

“…Besides, collection and archiving of frozen tissue for IFT is difficult, but it is easy to perform retrospective studies on formalin fixed paraffin embedded tissues by immunoperoxidase technique. IHC techniques for demonstration of PI-3 virus in tissues from animals with respiratory disease can be rapid and reliable; however the accomplishment of this procedure is dependent upon fixation techniques and quality of antisera (Haines et al, 1992). In this investigation, polyclonal rabbit-bovine anti-PI-3 virus serum was used for IHC detection of PI-3 virus antigens in cases of naturally occurring caprine pneumonia.…”

“…In ultrastructural examinations, the replication of PI-3 virus has been observed within epithelial cells of the respiratory tract and within alveolar macrophages (Bryson et al, 1983). The immunostaining with polyclonal antibodies in calves experimentally infected with PI-3 virus has demonstrated diffuse cytoplasmic staining with accentuation of the alveolar and bronchial epithelial cell surfaces (Haines et al, 1992). In rats experimentally infected with bovine PI-3 virus, viral antigens were present in the bronchial, bronchiolar and alveolar epithelial cells (Breker-Klassen et al, 1995).…”

“…Additional cross-sections from these samples, kept in paraffin blocks for 4 years, were obtained and stained only for immunohistochemistry to demonstrate the presence of PI-3 virus antigens on cases of 42 pneumonic lungs (except the verminous pneumonia). IHC staining was performed with the avidinbiotin-peroxidase complex (ABC) procedure (Haines et al, 1992), using commercially available immunoperoxidase kits (CadenzaTags peroxidase Kit, No: 407300; Shandon Inc., Pittsburgh, PA, USA). For immunohistochemistry, deparaffinized tissue sections were incubated with 3 H 2 O 2 in 70% methanol to inhibit endogenous peroxidase activity, and then washed three times in phosphate-buffered saline (PBS).…”

Recombinant type 5 adenoviruses expressing bovine parainfluenza virus type 3 glycoproteins protect Sigmodon hispidus cotton rats from bovine parainfluenza virus type 3 infection

“…Production of rabbit anti-bPIV3 serum. New Zealand White rabbits were immunized with 250 g of 30 to 60% (wt/vol) sucrose gradient-purified bPIV3 and Freund's adjuvant as previously described (17).…”