Application of Western Blotting to Diagnosis of Viral Infections

Abstract

Molecular biological techniques have an increasing role in the laboratory diagnosis of viral infections as a consequence of their being more sensitive and specific than methods developed in the past. This chapter discusses the principle, describes the methodology, and provides some practical clinical applications of Western blotting. Enzyme immunoassays (EIA) often are used for diagnosis of viral diseases as well as for screening of blood and blood products for viruses. The basic approach to the use of Western blotting for the diagnosis of viral infections begins with purified virions that are disrupted by ionic detergent treatment, releasing viral proteins. Chemiluminescence has two main advantages compared to chromogenic techniques. First, it enables a >10-fold increase in sensitivity without the use of isotopes, and second, exposure times can be varied to increase or decrease sensitivity. A potential competitor for the Western blot assay has been reported for the diagnosis of HIV infection. This new technique, called “recombinant-antigen immunoblot assay” (RIBA-HIV216), utilizes a set of purified antigens produced by recombinant technology. It remains to be determined whether this recombinant-protein assay is more specific and/or more sensitive than the standard Western blot assay. Nitrocellulose strips with blotted viral proteins or complete kits with all necessary reagents are available from commercial sources for human immunodeficiency virus (HIV-1 ), human T-cell leukemia virus type 1 (HTLV-l), and hepatitis C virus Western blot assays.