Single Molecule Detection of Fluorescently Labelled DNA Dendrimers

Abstract Single molecule detection has been achieved via many highly sophisticated microscopic techniques. Here we describe the detection of single molecules with conventional epifluorescent microscopy. The key to the technique is the use of DNA dendrimers DNA dendrimers have demonstrated utility in nucleic acid blots, Southerns, Northerns, etc. Typically DNA dendrimers yield 50-100 fold gain in signal over comparably labeled oligonucleotides. Immunodendrimers, DNA dendrimers conjugated to antibody molecules, have also been constructed and utilized in western blot assays. Individual, i.e. single molecule, 4- layer dendrimers, are readily detectable as point sources via conventional fluorescence microscopy and are useful for in situ hybridization and flow fluorescence quantitation. Nucleic acid hybridization is the underlying principle behind DNA dendrimer assembly. The “monomer” of DNA dendrimers consists of partially double stranded heteroduplexed DNA. Each monomer has an approximately 50 base double stranded “waist” surrounded by four approximately 30 base single stranded “arms”.