Date of Award:

5-1-1996

Document Type:

Thesis

Degree Name:

Master of Science (MS)

Department:

Biology

Committee Chair(s)

Bill B. Barnett

Committee

Bill B. Barnett

Committee

Reed P. Warren

Committee

Robert W. Sidwell

Abstract

Human cytomegalovirus (HCMV) is an important pathogen producing a wide spectrum of clinical syndromes in immunocompromised patients. HCMV is the most serious opportunistic viral pathogen in AIDS and organ transplant recipients. Once infected, an individual may carry the virus for a lifetime. With diminished immune status, HCMV reactivation often occurs. Murine cytomegalovirus (MCMV) infection in severe combined immunodeficient (SCID) mice is a model for HCMV infection in immunocompromised patients. The SCID mouse model responds to antiviral therapy in a manner similar to HCMV infections in immunodeficient humans. A panel of monoclonal antibodies (MAbs) towards MCMV was generated. Monoclonal antibodies and polyclonal antibodies raised in mice were characterized by their virus-neutralizing activity with or without complement. MCMV was neutralized by the MAbs both prior to virus adsorption to the cells and after virus adsorption to cells. The polyclonal antiserum produced was specific for MCMV with or without complement. The C' dependency of individual MAbs was an all-or-nothing phenomenon. A cytopathic effect, or CPE-based endpoint dilution assay in 96-well plates, was developed as a method for quantifying infectious MCMV. The effects of centrifugation of the inoculum onto cells, length of infection, the multiplicity of infection, and other parameters on MCMV yields were studied and standardized to an optimal assay method. Human plasma samples functioned as well or better than rabbit C' in satisfying the C' dependency of murine monoclonal antibodies in neutralizing MCMV. The effects of various whole and depleted complement sources on MCMV neutralization by C'-dependent MAbs were studied. The alternative pathway was not required for neutralization of MCMV by the complement-dependent antibody. The neutralization proceeded through the classical pathway.

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