CHEMILUMINESCENT WESTERN BLOT SYSTEM FOR AIDS DIAGNOSIS

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The goal of Phase I of this proposal is to establish a chemiluminescent detection system for AIDS Western blotting based on the chemiluminescent substrate for alkaline phosphatase, AMPPD (trademark). One advantage of this system will be increased sensitivity, over existing Western blots tests, which result in stained membrane strips. Also, a permanent record of results will be produced directly on film. A model Western blot system using bovine serum albumin (BSA) and commercially available antibodies will be used to develop a new luminescence amplifying system which will be necessary for nitrocellulose membrane. Blocking conditions and sensitivity will also be optimized. The luminescent amplifying system will result in a chemiluminescent emission with a maximum at approximately 540 nm. This wavelength is a good match for capturing the chemiluminescent signal on Polaroid instant black and white film. The optimal conditions developed with the BSA model system will then be applied to components of a DuPont HIV Western blot kit, using the kit's noninfectious controls to test the performance and sensitivity of the chemiluminescent system. The results obtained on instant film will be suitable for