Nucleic Acids Research, Vol 27, Issue 23 4553-4561, Copyright © 1999 by Oxford University Press
CM Niemeyer, M Adler, B Pignataro, S Lenhert, S Gao, L Chi, H Fuchs and D Blohm
The self-assembly of bis-biotinylated double-stranded DNA and the
tetravalent biotin-binding protein streptavidin (STV) have been studied by
non-denaturing gel electrophoresis and atomic force microscopy (AFM). The
rapid self-assembly reproducibly generated populations of individual
oligomeric complexes. Most strikingly, the oligomers predominantly
contained bivalent STV molecules bridging two adjacent DNA fragments to
form linear nanostructures. Trivalent STV branch points occurred with a
lower frequency and the presence of tetravalent STV was scarce. However,
valency distribution, size and the exchange dynamics of the supramolecular
aggregates were highly sensitive to stoichiometric variations in the
relative molar coupling ratio of bis- biotinylated DNA and STV. The largest
aggregates were obtained from equimolar amounts while excess STV led to the
formation of smaller oligomers appearing as fingerprint-like band patterns
in electrophoresis. Excess DNA, however, induces a complete breakdown of
the oligomers, likely a consequence of the instability of STV conjugates
containing more than two biotinylated DNA fragments. It was demonstrated
that the oligomers can further be functionalized, for instance by the
coupling of biotinylated immunoglobulins. Both pure and also
antibody-modified DNA-STV oligomers were used as reagents in immuno-PCR
(IPCR), a highly sensitive detection method for proteins and other
antigens. Employment of the supramolecular reagents led to an approximately
100-fold enhanced sensitivity compared to the conventional IPCR procedure.
ARTICLES
Self-assembly of DNA-streptavidin nanostructures and their use as reagents in immuno-PCR
Universitat Bremen, FB 2-UFT, Biotechnologie und Molekulare Genetik, Leobener Strasse, D-28359 Bremen, Germany. cmn@biotec.uni-bremen.de
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