The long-term objectives of this research are to develop a better understanding of the molecular events involved in cytomegalovirus (CMV) replication, and to contribute to the development of strategies for diagnosing and treating CMV-related diseases of man. Attention will be focused on three sets of CMV proteins that results obtained during the first grant period have indicated are both central to the virus' replication cycle, and of interest in a clinical diagnostic and therapeutic context. The first of these are DNA-binding proteins and are expressed early during virus replication. The second set consists of structural phosphoproteins of the virus particle, which are abundant and appear to elicit a strong and lasting immune response in both man and laboratory animals. And the third set comprises the viral glycoproteins, which are of interest in terms of their biogenesis and function, and of importance as subunit vaccine candidates.
The specific aims are to (i) prepare antibodies (monoclonal and polyclonal) to these proteins; (ii) use resulting antibodies to study the corresponding protein's function; (iii) adapt immunological reagents to diagnostic assays and compare their usefulness for this purpose with that of nucleic acid probes; and (iv) use the lamda gtll expression vector system, in conjunction with antibodies mentioned above to identify, map and study the CMV genes coding for proteins of interest. Procedures used will include protein purification; poly- and monoclonal antibody production; antibody assays by immunofluorescence, ELISA, immunoprecipitation and """"""""Towbin gel replica immunoassay""""""""; infectivity analyses; and recombinant DNA methodologies for producing, propagating and characterizing specific fragments of the CMV genome.
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