Accurate single-day titration of adenovirus vectors based on equivalence of protein VII nuclear dots and infectious particles.
Protein VII is an abundant component of adenovirus particles and is tightly associated with the viral DNA. It enters the nucleus along with the infecting viral genome and remains bound throughout early phase. Protein VII can be visualized by immunofluorescent staining as discrete dots in the infected cell nucleus. Comparison between protein VII staining and expression of the 72kDa DNA-binding protein revealed a one-to-one correspondence between protein VII dots and infectious viral genomes. A similar relationship was observed for a helper-dependent adenovirus vector expressing green fluorescent protein. This relationship allowed accurate titration of adenovirus preparations, including wild-type and helper-dependent vectors, using a 1-day immunofluorescence method. The method can be applied to any adenovirus vector and gives results equivalent to the standard plaque assay.
Walkiewicz
Marcin P
MP
Department of Microbiology, University of Virginia Health System, P.O. Box 800734, Charlottesville, VA 22908-0734, USA. mw3jb@virginia.edu
Morral
Nuria
N
Engel
Daniel A
DA
D000256:Adenoviridae / Q000737:chemistry* / Q000302:isolation & purification*; D004279:DNA, Viral / Q000032:analysis* / Q000235:genetics; D005455:Fluorescent Antibody Technique; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D014758:Viral Core Proteins / Q000032:analysis* / Q000276:immunology; D010948:Viral Plaque Assay
D004279:DNA, Viral; D014758:Viral Core Proteins; D049452:Green Fluorescent Proteins