In basic and applied HIV research, reliable detection of viral components is crucial to monitor progression of infection. While it is routine to detect structural viral proteins in vitro for diagnostic purposes, it was previously impossible to directly and dynamically visualize HIV in living cells without genetic modification of the virus.
This paper describes a novel fluorescent biosensor to dynamically trace HIV-1 morphogenesis in living cells. A single domain antibody that specifically binds the HIV-1 capsid protein (CA) at subnanomolar affinity was fused it to fluorescent proteins. The resulting fluorescent chromobody specifically recognizes the CA-harbouring HIV-1 Gag precursor protein in living cells and is applicable in various advanced light microscopy systems.
Confocal live cell microscopy and super-resolution microscopy allowed detection and dynamic tracing of individual virion assemblies at the plasma membrane. The analysis of subcellular binding kinetics showed cytoplasmic antigen recognition and incorporation into virion assembly sites. This new reporter system was used for automated image analysis, providing a new tool for cell-based HIV research. Generation of additional chromobodies with different binding sites should enable kinetic and functional studies to specifically probe the HIV assembly process.