Structure and DNA Hybridization Properties of Mixed Nucleic Acid/Maleimide-Ethylene Glycol Monolayers
The surface structure and DNA hybridization performance of thiolated single-strand DNA (HS-ssDNA) covalently attached to a maleimide-ethylene glycol disulfide (MEG) monolayer on gold have been investigated. Monolayer immobilization chemistry and surface coverage of reactive ssDNA probes were studied by X-ray photoelectron spectroscopy and time-of-flight secondary ion mass spectrometry. Orientation of the ssDNA probes was determined by near-edge X-ray absorption fine structure (NEXAFS). Target DNA hybridization on the DNA-MEG probe surfaces was measured by surface plasmon resonance (SPR) to demonstrate the utility of these probe surfaces for detection of DNA targets from both purified target DNA samples and complex biological mixtures such as blood serum. Data from complementary techniques showed that immobilized ssDNA density is strongly dependent on the spotted bulk DNA concentration and buffer ionic strength. Variation of the immobilized ssDNA density had a profound influence on the DNA probe orientation at the surface and subsequent target hybridization efficiency. With increasing surface probe density, NEXAFS polarization dependence results (followed by monitoring the N 1s {yields} {pi}* transition) indicate that the immobilized ssDNA molecules reorient toward a more upright position on the MEG monolayer. SPR assays of DNA targets from buffer and serum showed that DNA hybridization efficiency increased with decreasing surface probe density. However, target detection in serum was better on the 'high-density' probe surface than on the 'high-efficiency' probe surface. The amounts of target detected for both ssDNA surfaces were several orders of magnitude poorer in serum than in purified DNA samples due to nonspecific serum protein adsorption onto the sensing surface.
- Research Organization:
- Brookhaven National Lab. (BNL), Upton, NY (United States). National Synchrotron Light Source
- Sponsoring Organization:
- Doe - Office Of Science
- DOE Contract Number:
- DE-AC02-98CH10886
- OSTI ID:
- 930464
- Report Number(s):
- BNL-81216-2008-JA; ANCHAM; TRN: US0901395
- Journal Information:
- Analytical Chemistry, Vol. 79, Issue 12; ISSN 0003-2700
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
72 PHYSICS OF ELEMENTARY PARTICLES AND FIELDS
ABSORPTION
ADSORPTION
BLOOD SERUM
BUFFERS
CHEMISTRY
DETECTION
DISULFIDES
DNA
DNA HYBRIDIZATION
EFFICIENCY
FINE STRUCTURE
GLYCOLS
GOLD
HYBRIDIZATION
MASS SPECTROSCOPY
MIXTURES
MONITORING
ORIENTATION
PERFORMANCE
PLASMONS
POLARIZATION
PROBES
PROTEINS
RESONANCE
X-RAY PHOTOELECTRON SPECTROSCOPY
national synchrotron light source