PREPARATION OF ENVELOPE GLYCOPROTEIN RICH HIV-1

HIV-1 ENVELOPE GLYCOPROTEINS (GP) ARE VITAL COMPONENTS OF VIRAL PREPARATIONS USED IN SEROLOGICAL ASSAYS. THEY ARE ALSO CRITICAL FOR VIRAL RECOGNITION OF TARGET CELLS. CONSEQUENTLY, PURIFIED VIRUS WITH ENHANCED GP MAY HAVE INCREASED DIAGNOSTIC VALUE. VIRIONS WITH INCREASED GLYCOPROTEIN CONTENT MAY BE MORE SENSITIVE FOR NEUTRALIZATION ASSAYS. CHRONICALLY INFECTED, RETROVIRUS PRODUCING CELL LINES OFTEN YIELD REDUCED QUANTITIES OF VIRAL ENVELOPE GLYCOPROTEIN (GP). THIS REDUCED YIELD DEPENDS NOT ONLY ON THE CELL LINE USED BUT ALSO UPON SHEARING OF GLYCOPROTEINS DURING GRADIENT PURIFICATION. CLONES OF AN HIV-1 PRODUCING CELL LINE THAT ARE RICH IN SECRETED ENVELOPE GP HAVE BEEN REPORTED. BECAUSE CHRONIC CELL LINES ARE MORE MANAGEABLE AND THEIR PERFORMANCE MORE REPRODUCIBLE, THEY ARE PREFERRED OVER THE MORE CUMBERSOME, LESS RELIABLE LYTIC SYSTEMS. CHRONIC INFECTIONS WILL BE ESTABLISHED IN CLONES OF HUT 78 USING TWO SELECTED STRAINS OF HIV-1. PREPARED SUPERNATANTS FROM THESE CLONE CULTURES WILL BE USED FOR VIRUS PURIFICATION. A TWO PHASE AQUEOUS EXTRACTION SYSTEM HAS BEEN USED TO PURIFY FELINE LEUKEMIA VIRUS (FELV), A RELATED RETROVIRUS. VIRUSES RECOVERED FROM THAT SYSTEM SHOWED SIGNIFICANT INCREASES IN GP CONTENT. THIS STUDY WILL EVALUATE EXTRACTION PROCESSES FOR HIV-1. THE PRODUCT WILL BE ASSESSED USING A GP IMMUNOASSAY, WESTERN BLOT, AND INFECTIVITY ASSAYS.