DNA-Modified Electrodes Fabricated Using Copper-Free
Click Chemistry for Enhanced Protein Detection
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Abstract
A method of DNA monolayer formation
has been developed using copper-free
click chemistry that yields enhanced surface homogeneity and enables
variation in the amount of DNA assembled; extremely low-density DNA
monolayers, with as little as 5% of the monolayer being DNA, have
been formed. These DNA-modified electrodes (DMEs) were characterized
visually, with AFM, and electrochemically, and were found to facilitate
DNA-mediated reduction of a distally bound redox probe. These low-density
monolayers were found to be more homogeneous than traditional thiol-modified
DNA monolayers, with greater helix accessibility through an increased
surface area-to-volume ratio. Protein binding efficiency of the transcriptional
activator TATA-binding protein (TBP) was also investigated on these
surfaces and compared to that on DNA monolayers formed with standard
thiol-modified DNA. Our low-density monolayers were found to be extremely
sensitive to TBP binding, with a signal decrease in excess of 75%
for 150 nM protein. This protein was detectable at 4 nM, on the order
of its dissociation constant, with our low-density monolayers. The
improved DNA helix accessibility and sensitivity of our low-density
DNA monolayers to TBP binding reflects the general utility of this
method of DNA monolayer formation for DNA-based electrochemical sensor
development